Systematic Evaluation of Benchmark G4 Probes and G4 Clinical Drugs using three Biophysical Methods: A Guideline to Evaluate Rapidly G4-Binding Affinity.

G-quadruplex DNA structures (G4) are proven to interfere with most genetic and epigenetic processes. Small molecules binding these structures (G4 ligands) are invaluable tools to probe G4-biology and address G4-druggability in various diseases (cancer, viral infections). However, the large number of reported G4 ligands (>1000) could lead to confusion while selecting one for a given application. Herein we conducted a systematic affinity ranking of 11 popular G4 ligands vs 5 classical G4 sequences using FRET-melting, G4-FID assays and SPR. Interestingly SPR data globally align with the rankings obtained from the two semi-quantitative assays despite discrepancies due to limits and characteristics of each assay. In the whole, PhenDC3 emerges as the most potent binder irrespective of the G4 sequence. Immediately below PDS, PDC-360A, BRACO19, TMPyP4 and RHPS4 feature strong to medium binding again with poor G4 topology discrimination. More strikingly, the G4 drugs Quarfloxin, CX5461 and c-PDS exhibit weak affinity with all G4s studied. Finally, NMM and Cu-ttpy showed heterogeneous behaviors due, in part, to their physicochemical particularities poorly compatible with screening conditions. The remarkable properties of PhenDC3 led us to propose its use for benchmarking FRET-melting and G4-FID assays for rapid G4-affinity evaluation of newly developed ligands.

A dynamical measure of the black hole mass in a quasar 11 billion years ago.

Tight relationships exist in the local Universe between the central stellar properties of galaxies and the mass of their supermassive black hole (SMBH)1-3. These suggest that galaxies and black holes co-evolve, with the main regulation mechanism being energetic feedback from accretion onto the black hole during its quasar phase4-6. A crucial question is how the relationship between black holes and galaxies evolves with time; a key epoch to examine this relationship is at the peaks of star formation and black hole growth 8-12 billion years ago (redshifts 1-3)7. Here we report a dynamical measurement of the mass of the black hole in a luminous quasar at a redshift of 2, with a look back in time of 11 billion years, by spatially resolving the broad-line region (BLR). We detect a 40-µas (0.31-pc) spatial offset between the red and blue photocentres of the Hα line that traces the velocity gradient of a rotating BLR. The flux and differential phase spectra are well reproduced by a thick, moderately inclined disk of gas clouds within the sphere of influence of a central black hole with a mass of 3.2 × 108 solar masses. Molecular gas data reveal a dynamical mass for the host galaxy of 6 × 1011 solar masses, which indicates an undermassive black hole accreting at a super-Eddington rate. This suggests a host galaxy that grew faster than the SMBH, indicating a delay between galaxy and black hole formation for some systems.

Negative SPR Signals during Low Molecular Weight Analyte Recognition.

Surface plasmon resonance (SPR) is a powerful technique for studying biomolecular interactions mainly due to its sensitivity and real-time and label free advantages. While SPR signals are usually positive, only a few studies have reported sensorgrams with negative signals. The aim of the present work is to investigate and to explain the observation of negative SPR signals with the hypothesis that it reflects major changes in ligand conformation resulting from target binding. We demonstrated that these negative unconventional signals were due to the negative complex (ligand/analyte) refractive index increment (RII) deviation from the sum of the RII of the individual entities which counterbalanced the theoretical increase of the signal triggered by the target recognition and the ligand folding. We also found that the conformation change of biomolecules can induce a negative or a positive complex RII deviation depending on its sequence and immobilization mode.

Surface plasmon resonance study of the interaction of N-methyl mesoporphyrin IX with G-quadruplex DNA.

Surface plasmon resonance (SPR) was used to investigate the interaction between N-methyl mesoporphyrin IX (NMM) and different G-quadruplex (G4) topologies. The study was associated with circular dichroism analysis (CD) to assess the topology of the G4s when they interacted with NMM. We demonstrate the high selectivity of NMM for the parallel G4 structure with a dissociation constant at least ten times lower than those of other G4 topologies. We also confirm the ability of NMM to shift the G4 conformation from both the hybrid and antiparallel topologies toward the parallel structure.

Influence of the SPR Experimental Conditions on the G-Quadruplex DNA Recognition by Porphyrin Derivatives.

Surface plasmon resonance (SPR) is a powerful technique to study the interactions of ligands with analytes and therefore a number of biosensor surfaces and injection methods have been developed so far. However, many experimental parameters can affect the interactions and consequently the affinity measurements. In particular, the interactions of positively charged analytes (often used for anionic nucleic acids targets) can be influenced by the sensing surfaces (e.g., negatively charged), leading to significant nonspecific interactions as well as regeneration problems. The aim of the present work is to investigate the effect of different parameters, including ionic strength, SPR biosensor (i.e., nature of the surfaces), and the injection method on the recognition of porphyrin G-quadruplex ligands. We demonstrate that the injection method does not influence the affinity whereas the ionic strength and the nature of the surface impact the recognition properties of the porphyrin for the G-quadruplex DNA. We also found that self-assembled monolayer coating surface presents many advantages in comparison with carboxymethylated dextran surface for SPR studies of G-quadruplex DNA/ligand interactions: (i) the electrostatic interaction with charged analytes is less important, (ii) its structure/composition is less sensitive to the ionic concentration and less prone to unspecific adsorption, (iii) it is easily homemade, and (iv) the cost is approximately 10 times cheaper.

[Granulomatous hepatitis revealing a Mycobacterium bovis widespread infection following intravesical BCG therapy]. / Hépatite granulomateuse révélant une infection disséminée à Mycobacterium bovis après BCG-thérapie intravésicale.

INTRODUCTION: Intravesical therapy with bacillus Calmette-Guérin (BCG) has proved to be effective in the treatment of superficial bladder tumors. Side-effects include local infections and rarely disseminated BCG infection with multiple end organ complications such as granulomatous hepatitis, pneumonitis, aortitis and bone marrow involvement. CASE REPORT: We report an 83-year-old man who presented with chronic granulomatous hepatitis. This was related to intravesical BCG therapy received two years earlier for superficial bladder cancer. Aortitis, splenic infarction and hematopoietic involvement were also diagnosed. Outcome was favorable following adapted antibiotic course. CONCLUSION: This case report highlights the possibility of widespread BCG infection following intravesical treatment, and the need for vigilance in patients with a history of such a therapy even several years later.

In vitro biocompatibility of a dentine substitute cement on human MG63 osteoblasts cells: Biodentine™ versus MTA(®).

AIM: To compare the in vitro biocompatibility of Biodentine™ and White ProRoot(®) mineral trioxide aggregate (MTA(®) ) with MG63 osteoblast-like cells and to characterize the cement surface. METHODOLOGY: A direct contact model for MG63 osteoblast-like cells with cements was used for 1, 3 and 5 days. Four end-points were investigated: (i) cement surface characterization by atomic force microscopy (AFM), (ii) cell viability by MTT assay, (iii) protein amount quantification by Bradford assay and (iv) cell morphology by SEM. Statistical analyses were performed by analysis of variance (anova) with a repetition test method. RESULTS: The roughness of the cements was comparable as revealed by AFM analysis. The MTT test for Biodentine™ was similar to that of MTA(®) . Biodentine™ and MTA(®) induced a similar but slight decrease in metabolic activity. The amount of total protein was significantly enhanced at day three (P < 0.05) but slightly decreased at day five for both tested samples. Biodentine™ was tolerated as well as MTA(®) in all cytotoxicity assays. SEM observations showed improvement of cell attachment and proliferation on both material surfaces following the three incubation periods. CONCLUSION: The biocompatibility of Biodentine™ to bone cells was comparable to MTA(®) .

Factors associated with effective reassessment of antibiotic therapy on day 3.

UNLABELLED: Reassessment of antibiotic therapy (RA) after 3 days is constitutive of French antibiotic stewardship. This delay is required because of the need for clinical reappraisal and for obtaining microbiological data. Our aim was to determine the factors associated with an effective RA. PATIENTS AND METHOD: A prospective study was made in a 350-bed general hospital in which all prescriptions are computerized and validated daily by prescribers. All curative antibiotic therapies were reassessed during 4 weeks. RA was defined as effective if the initial antibiotic treatment was modified. All clinical, biological, and radiological data having contributed to the initial prescription and to RA were recorded during bedside visit with the prescribers, two hospital physicians and one infectious diseases specialist. RESULTS: In one month, 148 antibiotic treatments were reassessed. Pulmonary, digestive, and urinary infections accounted for two thirds of the cases. An effective RA was recorded in 28 cases (19%) and associated with hospitalization in the ICU (P=0.001), imaging supporting the diagnosis (P=0.016), and persistence or aggravation of clinical signs (P=0.007). Microbiological findings were not contributive to an effective RA. CONCLUSION: RA was associated to hospitalization in the ICU, to an inflammatory syndrome, and to the clinical outcome after 3 days. These results should help to improve the implementation of infectious diseases advice.

[Absence of interaction between thiomersal and aluminum occlusion chambers]. / Absence d'interaction entre le thiomersal et les chambres d'occlusion en aluminium.

BACKGROUND: Positive patch tests to thiomersal are frequent, but of little relevance. Interaction between aluminium occlusive chambers and the mercury of thiomersal may elicit skin irritation and cause false positive results. METHODS: Between January 1998 and May 1999, the patients were simultaneously patch tested with aluminium and plastic chambers. Positive skin tests were compared. RESULTS: Seventeen of 213 subjects showed positive tests: 15 with the 2 materials, 1 only for the aluminium chamber, 1 for the plastic chamber. DISCUSSION: The results did not differ for the two materials. We concluded that positive reactions observed were not caused by interaction between aluminium and thiomersal.

p73-deficient mice have neurological, pheromonal and inflammatory defects but lack spontaneous tumours.

p73 (ref. 1) has high homology with the tumour suppressor p53 (refs 2-4), as well as with p63, a gene implicated in the maintenance of epithelial stem cells. Despite the localization of the p73 gene to chromosome 1p36.3, a region of frequent aberration in a wide range of human cancers, and the ability of p73 to transactivate p53 target genes, it is unclear whether p73 functions as a tumour suppressor. Here we show that mice functionally deficient for all p73 isoforms exhibit profound defects, including hippocampal dysgenesis, hydrocephalus, chronic infections and inflammation, as well as abnormalities in pheromone sensory pathways. In contrast to p53-deficient mice, however, those lacking p73 show no increased susceptibility to spontaneous tumorigenesis. We report the mechanistic basis of the hippocampal dysgenesis and the loss of pheromone responses, and show that new, potentially dominant-negative, p73 variants are the predominant expression products of this gene in developing and adult tissues. Our data suggest that there is a marked divergence in the physiological functions of the p53 family members, and reveal unique roles for p73 in neurogenesis, sensory pathways and homeostatic control.

[Peritoneal fistulization in pleural aspergillosis associated with skin lesions]. / Fistulisation péritonéale d'une aspergillose pleurale associée à des lésions cutanées.

This report describes the development of a pleuroperitoneal fistula complicating pleural aspergillosis in a 63-year old non-immunocompromised man treated with itraconazole. The patient presented a confluent granuloma of the abdomen while an abscess of the abdominal wall was disclosed. Skin involvement, usually described in disseminated aspergillosis, has not been reported in pleuropulmonary aspergillosis.

Monoallelically expressed gene related to p53 at 1p36, a region frequently deleted in neuroblastoma and other human cancers.

We describe a gene encoding p73, a protein that shares considerable homology with the tumor suppressor p53. p73 maps to 1p36, a region frequently deleted in neuroblastoma and other tumors and thought to contain multiple tumor suppressor genes. Our analysis of neuroblastoma cell lines with 1p and p73 loss of heterozygosity failed to detect coding sequence mutations in remaining p73 alleles. However, the demonstration that p73 is monoallelically expressed supports the notion that it is a candidate gene in neuroblastoma. p73 also has the potential to activate p53 target genes and to interact with p53. We propose that the disregulation of p73 contributes to tumorigenesis and that p53-related proteins operate in a network of developmental and cell cycle controls.