Improved retroviral suicide gene transfer in colon cancer cell lines after cell synchronization with methotrexate

<p>Abstract</p> <p>Background</p> <p>Cancer gene therapy by retroviral vectors is mainly limited by the level of transduction. Retroviral gene transfer requires target cell division. Cell synchronization, obtained by drugs inducing a reversible inhibition of DNA synthesis, could therefore be proposed to precondition target cells to retroviral gene transfer. We tested whether drug-mediated cell synchronization could enhance the transfer efficiency of a retroviral-mediated gene encoding herpes simplex virus thymidine kinase (HSV-<it>tk</it>) in two colon cancer cell lines, DHDK12 and HT29.</p> <p>Methods</p> <p>Synchronization was induced by methotrexate (MTX), aracytin (ara-C) or aphidicolin. Gene transfer efficiency was assessed by the level of HSV-TK expression. Transduced cells were driven by ganciclovir (GCV) towards apoptosis that was assessed using annexin V labeling by quantitative flow cytometry.</p> <p>Results</p> <p>DHDK12 and HT29 cells were synchronized in S phase with MTX but not ara-C or aphidicolin. In synchronized DHDK12 and HT29 cells, the HSV-TK transduction rates were 2 and 1.5-fold higher than those obtained in control cells, respectively. Furthermore, the rate of apoptosis was increased two-fold in MTX-treated DHDK12 cells after treatment with GCV.</p> <p>Conclusions</p> <p>Our findings indicate that MTX-mediated synchronization of target cells allowed a significant improvement of retroviral HSV-<it>tk </it>gene transfer, resulting in an increased cell apoptosis in response to GCV. Pharmacological control of cell cycle may thus be a useful strategy to optimize the efficiency of retroviral-mediated cancer gene therapy.</p

Integrated immunovirological profiling validates plasma SARS-CoV-2 RNA as an early predictor of COVID-19 mortality.

peer reviewedDespite advances in COVID-19 management, identifying patients evolving toward death remains challenging. To identify early predictors of mortality within 60 days of symptom onset (DSO), we performed immunovirological assessments on plasma from 279 individuals. On samples collected at DSO11 in a discovery cohort, high severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral RNA (vRNA), low receptor binding domain–specific immunoglobulin G and antibody-dependent cellular cytotoxicity, and elevated cytokines and tissue injury markers were strongly associated with mortality, including in patients on mechanical ventilation. A three-variable model of vRNA, with predefined adjustment by age and sex, robustly identified patients with fatal outcome (adjusted hazard ratio for log-transformed vRNA = 3.5). This model remained robust in independent validation and confirmation cohorts. Since plasma vRNA’s predictive accuracy was maintained at earlier time points, its quantitation can help us understand disease heterogeneity and identify patients who may benefit from new therapies

Caractérisation des différences et des similarités entre carcinomes épidermoïdes et adénocarcinomes de l'oesophage (influence pronostique du type histologique)

LILLE2-BU Santé-Recherche (593502101) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

Le récepteur de l'EGF (médiateur de la surproduction de mucines et de l'inflammation dans l'épithélium biliaire. Implication dans la lithiase biliaire)

La lithiase biliaire, définie par la présence de calculs dans les voies biliaires, est un enjeu majeur de santé publique. Si la surproduction de mucines est un élément central de la lithogenèse biliaire, les mécanismes impliqués dans cette surproduction restent à identifier. Par une approche ex vivo et in vitro, notre travail a démontré que dans un contexte inflammatoire, l EGF-R, récepteur à activité tyrosine kinase, avait un rôle essentiel dans la surproduction de la mucine MUC5AC par les cellules épithéliales biliaires. Dans la seconde partie du travail, nous avons montré que, dans des cellules biliaires tumorales, le LPS active, une boucle positive de rétrocontrôle induisant l amplification de l activation de la voie de l EGF-R via la voie COX-2/PGE2. Cette amplification initiée par le LPS, présent dans la bile dans les pathologies biliaires inflammatoires, notamment lithiasiques, pourrait permettre de faire le lien entre l inflammation chronique et la carcinogenèse biliaire.PARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Esophageal Carcinoma: Prognostic Differences between Squamous Cell Carcinoma And Adenocarcinoma

International audienc

MUC5AC, a Gel-Forming Mucin Accumulating in Gallstone Disease, Is Overproduced via an Epidermal Growth Factor Receptor Pathway in the Human Gallbladder

Despite evidence that mucin overproduction is critical in the pathogenesis of gallstones, the mechanisms triggering mucin production in gallstone disease are unknown. Here, we tested the potential implication of an inflammation-dependent epidermal growth factor receptor (EGF-R) pathway in the regulation of gallbladder mucin synthesis. In gallbladder tissue sections from subjects with cholesterol gallstones, mucus accumulation was associated with neutrophil infiltration and with increased expressions of EGF-R and of tumor necrosis factor-α (TNF-α). In primary cultures of human gallbladder epithelial cells, TNF-α induced EGF-R overexpression. In the presence of TNF-α, EGF-R ligands (either EGF or transforming growth factor-α) caused significant increases in MUC5AC mRNA and protein production, whereas expression of the other gallbladder mucins MUC1, MUC3, and MUC5B was unchanged. In addition, on gallbladder tissue sections from subjects with gallstones, increased MUC5AC immunoreactivity was detected in the epithelium and within mucus gel in the lumen. Studies in primary cultures demonstrated that MUC5AC up-regulation induced by the combination of TNF-α with EGF-R ligands was completely blunted by inhibitors of EGF-R tyrosine kinase and mitogen-activated protein/extracellular signal-related kinase kinase. In conclusion, an inflammation-dependent EGF-R cascade causes overproduction of the gel-forming mucin MUC5AC, which accumulates in cholesterol gallstone disease. The ability to interrupt this cascade is of potential interest in the prevention of cholesterol gallstones

Recruitment of chromatin-modifying enzymes by CTIP2 promotes HIV-1 transcriptional silencing

Following entry and reverse transcription, the HIV-1 genome is integrated into the host genome. In contrast to productively infected cells, latently infected cells frequently harbor HIV-1 genomes integrated in heterochromatic structures, allowing persistence of transcriptionally silent proviruses. Microglial cells are the main HIV-1 target cells in the central nervous system and constitute an important reservoir for viral pathogenesis. In the present work, we show that, in microglial cells, the co-repressor COUP-TF interacting protein 2 (CTIP2) recruits a multienzymatic chromatin-modifying complex and establishes a heterochromatic environment at the HIV-1 promoter. We report that CTIP2 recruits histone deacetylase (HDAC)1 and HDAC2 to promote local histone H3 deacetylation at the HIV-1 promoter region. In addition, DNA-bound CTIP2 also associates with the histone methyltransferase SUV39H1, which increases local histone H3 lysine 9 methylation. This allows concomitant recruitment of HP1 proteins to the viral promoter and formation of local heterochromatin, leading to HIV-1 silencing. Altogether, our findings uncover new therapeutic opportunities for purging latent HIV-1 viruses from their cellular reservoirs