Perforated appendicitis due to fishbone.

Appendicitis is a common condition in daily clinical practice. Appendicitis due to foreign bodies is uncommon and may result from obstruction or perforation mechanism. We present a rare case of a 43-year-old male patient who was diagnosed with perforated appendicitis due to a fish bone by imaging studies and confirmed postoperatively. Confirming the fish bone causing the perforation on images is sometimes difficult, requiring the radiologist to actively search and determine the source. In addition to appendectomy, the surgeon also needs to pay attention to removing all foreign objects and treating perforations of surrounding organs.

Ligand-based discovery of new potential acetylcholinesterase inhibitors for Alzheimer's disease treatment.

The enzyme acetylcholinesterase (AChE) is currently a therapeutic target for the treatment of neurodegenerative diseases. These diseases have highly variable causes but irreversible evolutions. Although the treatments are palliative, they help relieve symptoms and allow a better quality of life, so the search for new therapeutic alternatives is the focus of many scientists worldwide. In this study, a QSAR-SVM classification model was developed by using the MATLAB numerical computation system and the molecular descriptors implemented in the Dragon software. The obtained parameters are adequate with accuracy of 88.63% for training set, 81.13% for cross-validation experiment and 81.15% for prediction set. In addition, its application domain was determined to guarantee the reliability of the predictions. Finally, the model was used to predict AChE inhibition by a group of quinazolinones and benzothiadiazine 1,1-dioxides obtained by chemical synthesis, resulting in 14 drug candidates with in silico activity comparable to acetylcholine.

Computational identification of chemical compounds with potential anti-Chagas activity using a classification tree.

Chagas disease is endemic to 21 Latin American countries and is a great public health problem in that region. Current chemotherapy remains unsatisfactory; consequently the need to search for new drugs persists. Here we present a new approach to identify novel compounds with potential anti-chagasic action. A large dataset of 584 compounds, obtained from the Drugs for Neglected Diseases initiative, was selected to develop the computational model. Dragon software was used to calculate the molecular descriptors and WEKA software to obtain the classification tree. The best model shows accuracy greater than 93.4% for the training set; the tree was also validated using a 10-fold cross-validation procedure and through a test set, achieving accuracy values over 90.5% and 92.2%, correspondingly. The values of sensitivity and specificity were around 90% in all series; also the false alarm rate values were under 10.5% for all sets. In addition, a simulated ligand-based virtual screening for several compounds recently reported as promising anti-chagasic agents was carried out, yielding good agreement between predictions and experimental results. Finally, the present work constitutes an example of how this rational computer-based method can help reduce the cost and increase the rate in which novel compounds are developed against Chagas disease.

Environmental sampling for SARS-CoV-2 at a reference laboratory and provincial hospital in central Viet Nam, 2020

@#Objective: To determine whether environmental surface contamination with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) occurred at a provincial hospital in Viet Nam that admitted patients with novel coronavirus disease 2019 (COVID-19) and at the regional reference laboratory responsible for confirmatory testing for SARS-CoV-2 in 2020. Methods: Environmental samples were collected from patient and staff areas at the hospital and various operational and staff areas at the laboratory. Specimens from frequently touched surfaces in all rooms were collected using a moistened swab rubbed over a 25 cm2 area for each surface. The swabs were immediately transported to the laboratory for testing by real-time reverse transcription polymerase chain reaction (RT-PCR). Throat specimens were collected from staff at both locations and were also tested for SARS-CoV-2 using real-time RT-PCR. Results: During the sampling period, the laboratory tested 6607 respiratory specimens for SARS-CoV-2 from patients within the region, and the hospital admitted 9 COVID-19 cases. Regular cleaning was conducted at both sites in accordance with infection prevention and control (IPC) practices. All 750 environmental samples (300 laboratory and 450 hospital) and 30 staff specimens were negative for SARS-CoV-2. Discussion: IPC measures at the facilities may have contributed to the negative results from the environmental samples. Other possible explanations include sampling late in a patient’s hospital stay when virus load was lower, having insufficient contact time with a surface or using insufficiently moist collection swabs. Further environmental sampling studies of SARS-CoV-2 should consider including testing for the environmental presence of viruses within laboratory settings, targeting the collection of samples to early in the course of a patient’s illness and including sampling of confirmed positive control surfaces, while maintaining appropriate biosafety measures.

Multiplexed blood-brain barrier organ-on-chip.

Organ-on-chip devices are intensively studied in academia and industry due to their high potential in pharmaceutical and biomedical applications. However, most of the existing organ-on-chip models focus on proof of concept of individual functional units without the possibility of testing multiple experimental stimuli in parallel. Here we developed a polydimethylsiloxane (PDMS) multiplexed chip with eight parallel channels branching from a common access port through which all eight channels can be addressed simultaneously without the need for extra pipetting steps thus increasing the reproducibility of the experimental results. At the same time, eight outlets provide individual entry to each channel with the opportunity to create eight different experimental conditions. A multiplexed chip can be assembled as a one-layer device for studying monocultures or as a two-layer device for studying barrier tissue functions. For a two-layer device, a ∼2 µm thick transparent PDMS membrane with 5 µm through-hole pores was fabricated in-house using a soft lithography technique, thereby allowing visual inspection of the cell-culture in real-time. The functionality of the chip was studied by recapitulating the blood-brain barrier. For this, human cerebral microvascular endothelial cells (hCMEC/D3) were cultured in mono- or coculture with human astrocytes. Immunostaining revealed a cellular monolayer with the expression of tight junction ZO-1 and adherence junction VE-cadherin proteins in endothelial cells as well as glial fibrillary acidic protein (GFAP) expression in astrocytes. Furthermore, multiplexed permeability studies of molecule passage through the cellular barrier exhibited expected high permeability coefficients for smaller molecules (4 kDa FITC-dextran) whereas larger molecules (20 kDa) crossed the barrier at a lower rate. With these results, we show that our device can be used as an organ-on-chip model for future multiplexed drug testing.

Machine learning-based models to predict modes of toxic action of phenols to Tetrahymena pyriformis.

The phenols are structurally heterogeneous pollutants and they present a variety of modes of toxic action (MOA), including polar narcotics, weak acid respiratory uncouplers, pro-electrophiles, and soft electrophiles. Because it is often difficult to determine correctly the mechanism of action of a compound, quantitative structure-activity relationship (QSAR) methods, which have proved their interest in toxicity prediction, can be used. In this work, several QSAR models for the prediction of MOA of 221 phenols to the ciliated protozoan Tetrahymena pyriformis, using Chemistry Development Kit descriptors, are reported. Four machine learning techniques (ML), k-nearest neighbours, support vector machine, classification trees, and artificial neural networks, have been used to develop several models with higher accuracies and predictive capabilities for distinguishing between four MOAs. They showed global accuracy values between 95.9% and 97.7% and area under Receiver Operator Curve values between 0.978 and 0.998; additionally, false alarm rate values were below 8.2% for training set. In order to validate our models, cross-validation (10-folds-out) and external test-set were performed with good behaviour in all cases. These models, obtained with ML techniques, were compared with others previously reported by other researchers, and the improvement was significant.

Phenotypic Heterogeneity in Attachment of Marine Bacteria toward Antifouling Copolymers Unraveled by AFM.

Up to recent years, bacterial adhesion has mostly been evaluated at the population level. Single cell level has improved in the past few years allowing a better comprehension of the implication of individual behaviors as compared to the one of a whole community. A new approach using atomic force microscopy (AFM) to measure adhesion forces between a live bacterium attached via a silica microbead to the AFM tipless cantilever and the surface has been recently developed. The objectives of this study is to examine the bacterial adhesion to a surface dedicated to ship hulls at the population and the cellular level to understand to what extent these two levels could be correlated. Adhesion of marine bacteria on inert surfaces are poorly studied in particular when substrata are dedicated to ship hulls. Studying these interactions in this context are worthwhile as they may involve different adhesion behaviors, taking place in salty conditions, using different surfaces than the ones usually utilized in the literacy. FRC (fouling release coatings)-SPC (self-polishing coatings) hybrids antifouling coatings have been used as substrata and are of particular interest for designing environmentally friendly surfaces, combining progressive surface erosion and low adhesion properties. In this study, a hybrid coating has been synthetized and used to study the adhesion of three marine bacteria, displaying different surface characteristics, using microplate assays associated with confocal scanning laser microscopy (CSLM) and AFM. This study shows that the bacterial strain that appeared to have the weakest adhesion and biofilm formation abilities when evaluated at the population level using microplates assays and CSLM, displayed stronger adhesion forces on the same surfaces at the single cell level using AFM. In addition, one of the strains tested which presented a strong ability to adhere and to form biofilm at the population level, displayed a heterogeneous phenotypic behavior at the single cell level. Therefore, these results suggest that the evaluation of adhesion at the population level cannot always be correlated with adhesion forces measured individually by AFM and that some bacteria are prone to phenotypic heterogeneity among their population.

Quantitative structure-activity relationship analysis and virtual screening studies for identifying HDAC2 inhibitors from known HDAC bioactive chemical libraries.

Histone deacetylases (HDAC) are emerging as promising targets in cancer, neuronal diseases and immune disorders. Computational modelling approaches have been widely applied for the virtual screening and rational design of novel HDAC inhibitors. In this study, different machine learning (ML) techniques were applied for the development of models that accurately discriminate HDAC2 inhibitors form non-inhibitors. The obtained models showed encouraging results, with the global accuracy in the external set ranging from 0.83 to 0.90. Various aspects related to the comparison of modelling techniques, applicability domain and descriptor interpretations were discussed. Finally, consensus predictions of these models were used for screening HDAC2 inhibitors from four chemical libraries whose bioactivities against HDAC1, HDAC3, HDAC6 and HDAC8 have been known. According to the results of virtual screening assays, structures of some hits with pair-isoform-selective activity (between HDAC2 and other HDACs) were revealed. This study illustrates the power of ML-based QSAR approaches for the screening and discovery of potent, isoform-selective HDACIs.

The acute promyelocytic leukaemia success story: curing leukaemia through targeted therapies.

The recent finding that almost all patients with acute promyelocytic leukaemia (APL) may be cured using a combination of retinoic acid (RA) and arsenic trioxide (As(2)O(3)) (N Engl J Med, 369, 2013 and 111) highlights the progress made in our understanding of APL pathogenesis and therapeutic approaches over the past 25 years. The study of APL has revealed many important lessons related to transcriptional control, nuclear organization, epigenetics and the role of proteolysis in biological control. Even more important has been the clinical demonstration that molecularly targeted therapy can eradicate disease.

Long-term survival of advanced triple-negative breast cancers with a dose-intense cyclophosphamide/anthracycline neoadjuvant regimen.

BACKGROUND: Triple-negative (TN) breast cancers exhibit major initial responses to neoadjuvant chemotherapy, but generally have a poor outcome. Because of the lack of validated drug targets, chemotherapy remains an important therapeutic tool in these cancers. METHODS: We report the survival of two consecutive series of 267 locally advanced breast cancers (LABC) treated with two different neoadjuvant regimens, either a dose-dense and dose-intense cyclophosphamide-anthracycline (AC) association (historically called SIM) or a conventional sequential association of cyclophosphamide and anthracycline, followed by taxanes (EC-T). We compared pathological responses and survival rates of these two groups and studied their association with tumours features. RESULTS: Although the two regimens showed equivalent pathological complete response (pCR) in the whole population (16 and 12%), the SIM regimen yielded a non-statistically higher pCR rate than EC-T (48% vs 24%, P=0.087) in TN tumours. In the SIM protocol, DFS was statistically higher for TN than for non-TN patients (P=0.019), although we showed that the TN status was associated with an increased initial risk of recurrence in both regimens. This effect gradually decreased and after 2 years, TN was associated with a significantly decreased likelihood of relapse in SIM-treated LABC (hazard ratio (HR)=0.25 (95% CI: 0.07-0.86), P=0.028). CONCLUSIONS: AC dose intensification treatment is associated with a very favourable long-term survival rate in TN breast cancers. These observations call for a prospective assessment of such dose-intense AC-based regimens in locally advanced TN tumours.

QSPR in oral bioavailability: specificity or integrality?

During the last decade the technological advances in drug discovery changed the absorption, distribution, metabolism, excretion and toxicity (ADMET) profiles of New Chemical Entities (NCEs). Among ADMET processes, absorption plays an important role in the research and development of more effective orally administered drugs. Although significant progress has been made in in vitro, in situ and in vivo experimental determinations of absorption, the development of in silico methodologies has emerged as a cheaper and fast alternative to predict them. Even though several in silico models have been described in the literature to predict oral bioavailability and related properties, the prediction accuracy and their potential use is still limited. The low precision and high variability of data, the lack of a complete experimental and theoretical validation of in silico approach, and above all, the multi-factorial nature of the oral absorption term, make the development of predictive in silico models a thorny task. The present review discusses several important advances regarding the QSPR approaches used in the development of predictive oral bioavailability models. The importance of fixing the problem associated with data resource, as well as improving the reliability of in silico results is highlighted. Optimization of individual properties along the absorption process must be integrated in a multi-objective scenario for studying oral bioavailability behavior in the early drug discovery and development.

Valproic acid induces differentiation and transient tumor regression, but spares leukemia-initiating activity in mouse models of APL.

Aberrant histone acetylation was physiopathologically associated with the development of acute myeloid leukemias (AMLs). Reversal of histone deacetylation by histone deacetylase inhibitor (HDACis) activates a cell death program that allows tumor regression in mouse models of AMLs. We have used several models of PML-RARA-driven acute promyelocytic leukemias (APLs) to analyze the in vivo effects of valproic acid, a well-characterized HDACis. Valproic acid (VPA)-induced rapid tumor regression and sharply prolonged survival. However, discontinuation of treatment was associated to an immediate relapse. In vivo, as well as ex vivo, VPA-induced terminal granulocytic differentiation. Yet, despite full differentiation, leukemia-initiating cell (LIC) activity was actually enhanced by VPA treatment. In contrast to all-trans retinoic acid (ATRA) or arsenic, VPA did not degrade PML-RARA. However, in combination with ATRA, VPA synergized for PML-RARA degradation and LIC eradication in vivo. Our studies indicate that VPA triggers differentiation, but spares LIC activity, further uncouple differentiation from APL clearance and stress the importance of PML-RARA degradation in APL cure.

(+)α-Tocopheryl succinate inhibits the mitochondrial respiratory chain complex I and is as effective as arsenic trioxide or ATRA against acute promyelocytic leukemia in vivo.

The vitamin E derivative (+)α-tocopheryl succinate (α-TOS) exerts pro-apoptotic effects in a wide range of tumors and is well tolerated by normal tissues. Previous studies point to a mitochondrial involvement in the action mechanism; however, the early steps have not been fully elucidated. In a model of acute promyelocytic leukemia (APL) derived from hCG-PML-RARα transgenic mice, we demonstrated that α-TOS is as effective as arsenic trioxide or all-trans retinoic acid, the current gold standards of therapy. We also demonstrated that α-TOS induces an early dissipation of the mitochondrial membrane potential in APL cells and studies with isolated mitochondria revealed that this action may result from the inhibition of mitochondrial respiratory chain complex I. Moreover, α-TOS promoted accumulation of reactive oxygen species hours before mitochondrial cytochrome c release and caspases activation. Therefore, an in vivo antileukemic action and a novel mitochondrial target were revealed for α-TOS, as well as mitochondrial respiratory complex I was highlighted as potential target for anticancer therapy.

A refined molecular taxonomy of breast cancer.

The current histoclinical breast cancer classification is simple but imprecise. Several molecular classifications of breast cancers based on expression profiling have been proposed as alternatives. However, their reliability and clinical utility have been repeatedly questioned, notably because most of them were derived from relatively small initial patient populations. We analyzed the transcriptomes of 537 breast tumors using three unsupervised classification methods. A core subset of 355 tumors was assigned to six clusters by all three methods. These six subgroups overlapped with previously defined molecular classes of breast cancer, but also showed important differences, notably the absence of an ERBB2 subgroup and the division of the large luminal ER+ group into four subgroups, two of them being highly proliferative. Of the six subgroups, four were ER+/PR+/AR+, one was ER-/PR-/AR+ and one was triple negative (AR-/ER-/PR-). ERBB2-amplified tumors were split between the ER-/PR-/AR+ subgroup and the highly proliferative ER+ LumC subgroup. Importantly, each of these six molecular subgroups showed specific copy-number alterations. Gene expression changes were correlated to specific signaling pathways. Each of these six subgroups showed very significant differences in tumor grade, metastatic sites, relapse-free survival or response to chemotherapy. All these findings were validated on large external datasets including more than 3000 tumors. Our data thus indicate that these six molecular subgroups represent well-defined clinico-biological entities of breast cancer. Their identification should facilitate the detection of novel prognostic factors or therapeutical targets in breast cancer.

Immunohistochemical and molecular analyses of HER2 status in breast cancers are highly concordant and complementary approaches.

BACKGROUND: Immunohistochemistry (IHC) and fluorescent in situ hybridisation (FISH) are currently the most commonly used methods to assess HER2 status. PCR-based assays allow quantitative determination of HER2 amplification (Q-PCR) or overexpression (Q-RT-PCR), but are not routinely used. We evaluated the relevance of Q-RT-PCR for HER2 status determination. METHODS: We compared IHC and Q-RT-PCR in 466 breast tumours. In discordant or equivocal cases, five additional methods (IHC with two other antibodies, FISH, silver in situ hybridisation (SISH) and Q-PCR) were combined to determine HER2 status. Two cases with HER2 intra-tumour heterogeneity were further explored by allelic profiles analysis and HUMARA clonality determination after microdissection. RESULTS: We observed 97.3% concordance between Q-RT-PCR and non-equivocal IHC. Twelve out of 466 cases (3%) revealed discordances between the two methods. The power of Q-RT-PCR to predict HER2 status (defined by seven methods) was similar to that of IHC. Although rare, some discordances between techniques might be due to HER2 intra-tumour heterogeneity and we report two examples, one tumour containing two distinct clones, another tumour consisting of HER2 amplified and non-amplified subclones. CONCLUSION: Q-RT-PCR and IHC are highly concordant methods for HER2 status assessment, and Q-RT-PCR allows a highly reliable quantitative assessment and could be a useful adjunct to IHC.

The tumor suppressor protein PML controls apoptosis induced by the HIV-1 envelope.

Promyelomonocytic leukemia (PML) is a prominent oncosuppressor whose inactivation is involved in the pathogenesis of hematological and epithelial cancers. Here, we report that PML aggregated in nuclear bodies in syncytia elicited by the envelope glycoprotein complex (Env) of human immunodeficiency virus-1 (HIV-1) in vitro. PML aggregation occurred after the fusion of nuclei (karyogamy) within syncytia but before the apoptotic program was activated. The aggregation of PML was detectable in syncytia present in the brain or lymph nodes from patients with HIV-1 infection, as well as in a fraction of blood leukocytes, correlating with viral status. Using a range of specific inhibitors of PML (the oncogenic PML/RARalpha fusion product or specific small interfering RNAs), we demonstrated that, in Env-elicited syncytia, PML was required for activating phosphorylation of ataxia telangiectasia mutated (ATM), which colocalized with PML in nuclear bodies, in a molecular complex that also involved topoisomerase IIbeta-binding protein 1. PML knockdown thus inhibited the ATM-dependent DNA damage response that culminates in the activation of p53, p53-dependent transcription of pro-apoptotic genes and cell death. Infection of CD4-expressing cells with HIV-1 also induced syncytial apoptosis, which could be suppressed by inhibiting PML. Altogether, these data indicate that PML activation is a critical early event that participates in the apoptotic demise of HIV-1-elicited syncytia.

Arsenic trioxide inhibits ATRA-induced prostaglandin E2 and cyclooxygenase-1 in NB4 cells, a model of acute promyelocytic leukemia.

In acute promyelocytic leukemia (APL), all-trans retinoic acid (ATRA) triggers cell differentiation, while arsenic trioxide (As(2)O(3)) generates partial differentiation and apoptosis. Animal and human studies suggest that newly diagnosed APL patients can be cured using As(2)O(3) combined with ATRA. Cyclooxygenases are involved in prostaglandins and thromboxane synthesis. We have recently demonstrated that ATRA induces cyclooxygenase-1 (COX-1) expression and prostaglandin synthesis in NB4 cells and in blasts from patients with APL. In the present study we investigated the effect of ATRA and As(2)O(3) co-treatment on COX-1 expression and prostaglandin formation and tested the effect of the COX-1/COX-2 nonselective inhibitor indomethacin on cell differentiation. Arsenic treatment of NB4 cells resulted in a partial but significant reduction of ATRA-dependent induction of COX-1 expression and activity. Pretreatment of NB4 cells with indomethacin significantly impaired ATRA/As(2)O(3)-induced differentiation, as assessed by cell morphology, nitroblue tetrazolium test or CD11c expression. PGE(2) reversed the negative effect of indomethacin on differentiation of ATRA/As(2)O(3)-treated NB4 cells. In conclusion, COX-1 contributes to ATRA-dependent maturation of NB4 cells and is affected by As(2)O(3). These results also suggest that nonsteroidal antiinflammatory drugs should be avoided in APL patients treated with the combination of ATRA and As(2)O(3).

Ubiquitylated Tax targets and binds the IKK signalosome at the centrosome.

Constitutive activation of the NF-kappaB pathway by the Tax oncoprotein plays a crucial role in the proliferation and transformation of HTLV-I infected T lymphocytes. We have previously shown that Tax ubiquitylation on C-terminal lysines is critical for binding of Tax to IkappaB kinase (IKK) and its subsequent activation. Here, we report that ubiquitylated Tax is not associated with active cytosolic IKK subunits, but binds endogenous IKK-alpha, -beta, -gamma, targeting them to the centrosome. K63-ubiquitylated Tax colocalizes at the centrosome with IKK-gamma, while K48-ubiquitylated Tax is stabilized upon proteasome inhibition. Altogether, these results support a model in which K63-ubiquitylated Tax activates IKK in a centrosome-associated signalosome, leading to the production of Tax-free active cytoplasmic IKK. These observations highlight an unsuspected link between Tax-induced IKK activation and the centrosome.

Changes in allelic imbalances in locally advanced breast cancers after chemotherapy.

In advanced breast cancers, TP53 mutation is highly predictive of complete response to high-dose epirubicin/cyclophosphamide chemotherapy. In these tumours with an altered control of genomic stability, accumulation of chemotherapy-induced genetic alterations may contribute to cell death and account for complete response. To explore the effects of chemotherapy on stability of the tumour genome, allelic profiles were obtained from microdissected tumour samples before and after chemotherapy in 29 unresponsive breast cancers (9 with TP53 mutation). Ninety-four per cent allelic profiles remained unchanged after treatment. Interestingly, 11 profiles (6%) showed important changes after treatment; allelic imbalances significantly increased (four cases) or decreased (seven cases) after chemotherapy in three distinct experiments, two of which using laser microdissected tumour cells. These genetic changes were not linked to the TP53 status, but one tumour showed complete disappearance of TP53-mutated cells in the residual tumour after treatment. Altogether, these observations carry important implications for the clonal evolution of breast cancers treated with DNA-damaging agents, as they point both to the importance of tumour heterogeneity and chemotherapy-driven selection of subclones.