Main Issues for a Fully Predictive Plasma Spray Torch Model and Numerical Considerations

International audienc

Quantifying RNA modifications by mass spectrometry: a novel source of biomarkers in oncology

International audienceDespite significant progress in targeted therapies, cancer recurrence remains a major cause of mortality worldwide. Identification of accurate biomarkers, through molecular profiling in healthy and cancer patient samples, will improve diagnosis and promote personalized medicine. While genetic and epigenetic alterations of DNA are currently exploited as cancer biomarkers, their robustness is limited by tumor heterogeneity. Recently, cancer-associated changes in RNA marks have emerged as a promising source of diagnostic and prognostic biomarkers. RNA epigenetics (also known as epitranscriptomics) is an emerging field in which at least 150 chemical modifications in all types of RNA (mRNA, tRNA, lncRNA, rRNA, and microRNA) have been detected. These modifications fine-tune gene expression in both physiological and pathological processes. A growing number of studies have established links between specific modified nucleoside levels in solid/liquid biopsies, and cancer onset and progression. In this review, we highlight the potential role of epitranscriptomic markers in refining cancer diagnosis and/or prognosis. RNA modification patterns may contain important information for establishing an initial diagnosis, monitoring disease evolution, and predicting response to treatment. Furthermore, recent developments in mass spectrometry allow reliable quantification of RNA marks in solid biopsies and biological fluids. We discuss the great potential of mass spectrometry for identifying epitranscriptomic biomarker signatures in cancer diagnosis. While there are various methods to quantify modified nucleosides, most are unable to detect and quantify more than one type of RNA modification at a time. Mass spectrometry analyses, especially GC-MS/MS and LC-MS/MS, overcome this limitation and simultaneously detect modified nucleosides by multiple reaction monitoring. Indeed, several groups are currently validating mass spectrometry methods that quantify several nucleosides at one time in liquid biopsies. The challenge now is to exploit these powerful analytical tools to establish epitranscriptomic signatures that should open new perspectives in personalized medicine. This review summarizes the growing clinical field of analysis of RNA modifications and discusses pre-analytical and analytical approaches, focusing in particular on the development of new mass spectrometry tools and their clinical applications

Les personnes vivant avec le VIH sont-elles plus confrontées à des seconds cancers que la population générale : travail issu du réseau CANCERVIH

International audienceIntroductionPeople living with HIV (PLWHIV) are at a higher risk of cancer compared to the general population. With improved cancer treatments and the increased life expectancy of PLWHIV, the incidence of second cancers is also expected to increase.MethodsWe reviewed the cases of PLWHIV with cancer that have been presented to the CANCERVIH national multidisciplinary board since 2014. We included all cases with a history of cancer, and studied the incidence and types of second cancers.ResultsIn total, 719 cases were reviewed, out of which 94 (13%) had a history of at least one cancer. For the first primary cancers, 46 (49%) were AIDS-defining cancers (ADCs) and 48 (51%) were non-AIDS-defining cancers (NADCs). Kaposi sarcoma (33%) and NHL (15%) occurred most frequently as first cancers. Among the first cancers that were ADCs, 15% of the second cancers were NHL, 11% anal canal cancers, 9% bladder and 9% Hodgkin lymphomas. Among the first cancers that were NADCs, 38% of the second cancers were lung cancers, 8% bladder, 8% head and neck and 8% NHL.DiscussionWith the aging of PLWHIV, the incidence of second and subsequent cancers is expected to increase in this population. Immuno-virological control should be maintained. Increased surveillance, early prevention and screening programs should be offered to all PLWHIV, including those with an undetectable HIV viral load and/or immune restoration.IntroductionLes personnes vivant avec le VIH (PVVIH) ont un risque plus élevé de cancer que la population générale. Avec l’amélioration des traitements carcinologiques et l’augmentation de l’espérance de vie des PVVIH, l’incidence des seconds cancers devrait augmenter.MéthodesNous avons examiné les dossiers des PVVIH atteintes d’un cancer qui ont été présentés en RCP nationale CANCERVIH depuis 2014. Nous avons inclus tous les cas avec antécédents de cancer, et étudié l’incidence et les types de seconds cancers.RésultatsAu total, 719 dossiers ont été examinés dont 94 (13 %) avaient un antécédent d’au moins un cancer. Pour les premiers cancers primitifs, 46 (49 %) étaient des cancers classant-SIDA (ADCs) et 48 (51 %) des cancers non-classant-SIDA (NADCs). Le sarcome de Kaposi (33 %) et le LNH (15 %) étaient les premiers cancers les plus fréquents. Parmi les premiers cancers qui étaient des ADCs, 15 % des seconds cancers étaient des LNH, 11 % des cancers du canal anal, 9 % de vessie et 9 % des lymphomes de Hodgkin. Parmi les premiers cancers qui étaient des NADCs, 38 % des seconds cancers étaient des cancers du poumon, 8 % de vessie, 8 % de la tête et cou et 8 % des LNH.DiscussionAvec le vieillissement des PVVIH, l’incidence des seconds cancers (et ultérieurs) devrait augmenter dans cette population. Le contrôle immuno-virologique doit être maintenu. Surveillance accrue, prévention précoce et programmes de dépistage devraient être proposés à toutes les PVVIH, même avec une charge virale VIH indétectable et/ou une restauration immunitaire

Antibiotics inhibit sphere-forming ability in suspension culture

International audienceBACKGROUND:This last decade, a lot of emphasis has been placed on developing new cancer cell culture models, closer to in vivo condition, in order to test new drugs and therapies. In the case of colorectal cancer, the use of patient biopsies to seed 3D primary cultures and mimic tumor initiation necessitates the use of antibiotics to prevent microbial intestinal contamination. However, not only long term use of antibiotics may mask the presence of low levels of microbial contamination, it may also impact cancer cell phenotype.METHODS:In this study we tested the impact of penicillin-streptomycin cocktail addition in both monolayer and suspension culture. To ensure the reliability of our observations we used six different cell lines and each experiment was performed in triplicate. Results were analyzed with Student's t test.RESULTS:We show that penicillin-streptomycin cocktail inhibits the sphere-forming ability of six cancer cell lines in suspension culture though it has no impact in monolayer culture. We correlate this effect with a significant decrease of cancer stem cells pool which holds self-renewal potential.CONCLUSIONS:Overall, this study warns against systematic addition of antibiotics in growth medium and raises the interesting possibility of using antibiotics to target cancer stem cells

The stability of Fbw7α in M-phase requires its phosphorylation by PKC

<div><p>Fbw7 is a tumor suppressor often deleted or mutated in human cancers. It serves as the substrate-recruiting subunit of a SCF ubiquitin ligase that targets numerous critical proteins for degradation, including oncoproteins and master transcription factors. Cyclin E was the first identified substrate of the SCF^Fbw7 ubiquitin ligase. In human cancers bearing <i>FBXW7</i>-gene mutations, deregulation of cyclin E turnover leads to its aberrant expression in mitosis. We investigated Fbw7 regulation in <i>Xenopus</i> eggs, which, although arrested in a mitotic-like phase, naturally express high levels of cyclin E. Here, we report that Fbw7α, the only Fbw7 isoform detected in eggs, is phosphorylated by PKC (protein kinase C) at a key residue (S18) in a manner coincident with Fbw7α inactivation. We show that this PKC-dependent phosphorylation and inactivation of Fbw7α also occurs in mitosis during human somatic cell cycles, and importantly is critical for Fbw7α stabilization itself upon nuclear envelope breakdown. Finally, we provide evidence that S18 phosphorylation, which lies within the intrinsically disordered N-terminal region specific to the α-isoform reduces the capacity of Fbw7α to dimerize and to bind cyclin E. Together, these findings implicate PKC in an evolutionarily-conserved pathway that aims to protect Fbw7α from degradation by keeping it transiently in a resting, inactive state.</p></div

Physical Modeling of Translation with a Ballistic Model: Application to the Estimation of the Kinetic Parameters from Ribosome Sequencing Experiments

International audienceGene expression consists in the synthesis of proteins from the information encoded on DNA. One of the two main steps of gene expression is the translation of messenger RNA (mRNA) into polypeptide sequences of amino acids. Here, by taking into account mRNA degradation, we model the motion of ribosomes along mRNA with a ballistic model where particles advance along a filament without excluded volume interactions. Unidirectional models of transport have previously been used to fit the average density of ribosomes obtained by the experimental ribosome sequencing (Ribo-Seq) technique. In this case an inverse fit gives access to the kinetic rates: the position-dependent speeds and the entry rate of ribosomes into mRNA. The degradation rate is not, however, accounted for and experimental data from differentexperiments are needed to have enough parameters for the fit. Here, we propose an entirely novel experimental setup and theoretical framework consisting in splitting the mRNAs into categories depending on the number of ribosomes from one to four. We analytically solve the ballistic model for a fixed number of ribosomes per mRNA, study the different regimes of degradation, and propose a criterion for the quality of the inverse fit. The proposed method provides a high sensitivity to the mRNA degradation rate. The additional equations coming from using the monosome (single ribosome) and polysome (arbitrary number) Ribo-Seq profiles enables us to determine all the kinetic rates in terms of the experimentally accessible mRNA degradation rate

DNA-methylation-dependent alterations of claudin-4 expression in human bladder carcinoma

International audienceThe expression pattern of tight junction (TJ) proteins is frequently disrupted in epithelial tumors. In particular, isoform- and organ-specific alterations of claudins have been detected in human cancers, highlighting them as interesting tools for the prognosis or treatment of various carcinomas. However, the molecular mechanisms responsible for these alterations are seldom identified. Here, we analyzed the expression and localization of claudins 1, 4, and 7 in human bladder carcinoma. Claudin-4 expression was significantly altered in 26/39 tumors, contrasting with the rare modifications detected in the expression of claudins 1 and 7. Overexpression of claudin-4 in differentiated carcinomas was followed by a strong downregulation in invasive/high-grade tumors, and this expression pattern was associated to the 1-year survival of bladder tumor patients. A CpG island was identified within the coding sequence of the CLDN4 gene, and treatment with a methyl-transferase inhibitor restored expression of the protein in primary cultures prepared from high-grade human bladder tumors. In addition, claudin-4 expression correlated with its gene methylation profile in healthy and tumoral bladders from 20 patients, and downregulation of claudin-4 expression was detected in the urothelium of mice overexpressing DNA methyl transferase 3a (Dnmt3a). Delocalization of claudins 1 and 4 from TJs was observed in most human bladder tumors and in the bladder tumor cell line HT-1376. Although the CLDN4 gene was unmethylated in these cells, pharmacological inhibition of methyl transferases re-addressed the two proteins to TJs, resulting in an increase of cell polarization and transepithelial resistance. These biological effects were prevented by expression of claudin-4-specific siRNAs, demonstrating the important role played by claudin-4 in maintaining a functional regulation of homeostasis in urothelial cells. Results of this study indicate that the TJ barrier is disrupted from early stages of urothelial tumorigenesis. In addition, we identified hypermethylation as the mechanism leading to the alteration of claudin-4 expression, and maybe also localization, in bladder carcinoma

Physical modeling of ribosomes along messenger RNA: estimating kinetic rates from ribosome profiling experiments with a ballistic model

Gene expression consists in the synthesis of proteins from the information encoded on DNA. One of the two main steps of gene expression is the translation of messenger RNA into polypeptide sequences of amino acids. Here, by taking into account messenger RNA degradation, we model the motion of ribosomes along messenger RNA with a ballistic model where particles advance along a filament without excluded volume interactions. Unidirectional models of transport have previously been used to fit the average density of ribosomes obtained by the experimental ribo-sequencing (Ribo-seq) technique. In this case an inverse fit gives access to the kinetic rates: the position-dependent speeds and the entry rate of ribosomes onto messenger RNA. The degradation rate is not, however, accounted for and experimental data from different experiments are needed to have enough parameters for the fit. Here, we propose an entirely novel experimental setup and theoretical framework consisting in splitting the messenger RNAs into categories depending on the number of ribosomes from one to four. We solve analytically the ballistic model for a fixed number of ribosomes per messenger RNA, study the different regimes of degradation, and propose a criteria for the quality of the inverse fit. The proposed method provides a high sensitivity to the messenger RNA degradation rate. The additional equations coming from using the monosome (single ribosome) and polysome (arbitrary number) ribo-seq profiles enable us to determine all the kinetic rates in terms of the experimentally accessible messenger RNA degradation rate