A Simplified Approach for the Molecular Classification of Glioblastomas

Glioblastoma (GBM) is the most common malignant primary brain tumors in adults and exhibit striking aggressiveness. Although GBM constitute a single histological entity, they exhibit considerable variability in biological behavior, resulting in significant differences in terms of prognosis and response to treatment. In an attempt to better understand the biology of GBM, many groups have performed high-scale profiling studies based on gene or protein expression. These studies have revealed the existence of several GBM subtypes. Although there remains to be a clear consensus, two to four major subtypes have been identified. Interestingly, these different subtypes are associated with both differential prognoses and responses to therapy. In the present study, we investigated an alternative immunohistochemistry (IHC)-based approach to achieve a molecular classification for GBM. For this purpose, a cohort of 100 surgical GBM samples was retrospectively evaluated by immunohistochemical analysis of EGFR, PDGFRA and p53. The quantitative analysis of these immunostainings allowed us to identify the following two GBM subtypes: the “Classical-like” (CL) subtype, characterized by EGFR-positive and p53- and PDGFRA-negative staining and the “Proneural-like” (PNL) subtype, characterized by p53- and/or PDGFRA-positive staining. This classification represents an independent prognostic factor in terms of overall survival compared to age, extent of resection and adjuvant treatment, with a significantly longer survival associated with the PNL subtype. Moreover, these two GBM subtypes exhibited different responses to chemotherapy. The addition of temozolomide to conventional radiotherapy significantly improved the survival of patients belonging to the CL subtype, but it did not affect the survival of patients belonging to the PNL subtype. We have thus shown that it is possible to differentiate between different clinically relevant subtypes of GBM by using IHC-based profiling, a method that is advantageous in its ease of daily implementation and in large-scale clinical application.     

Peer Review of Grant Applications: Criteria Used and Qualitative Study of Reviewer Practices

Background

Peer review of grant applications has been criticized as lacking reliability. Studies showing poor agreement among reviewers supported this possibility but usually focused on reviewers’ scores and failed to investigate reasons for disagreement. Here, our goal was to determine how reviewers rate applications, by investigating reviewer practices and grant assessment criteria.

Methods and Findings

We first collected and analyzed a convenience sample of French and international calls for proposals and assessment guidelines, from which we created an overall typology of assessment criteria comprising nine domains relevance to the call for proposals, usefulness, originality, innovativeness, methodology, feasibility, funding, ethical aspects, and writing of the grant application. We then performed a qualitative study of reviewer practices, particularly regarding the use of assessment criteria, among reviewers of the French Academic Hospital Research Grant Agencies (Programmes Hospitaliers de Recherche Clinique, PHRCs). Semi-structured interviews and observation sessions were conducted. Both the time spent assessing each grant application and the assessment methods varied across reviewers. The assessment criteria recommended by the PHRCs were listed by all reviewers as frequently evaluated and useful. However, use of the PHRC criteria was subjective and varied across reviewers. Some reviewers gave the same weight to each assessment criterion, whereas others considered originality to be the most important criterion (12/34), followed by methodology (10/34) and feasibility (4/34). Conceivably, this variability might adversely affect the reliability of the review process, and studies evaluating this hypothesis would be of interest.

Conclusions

Variability across reviewers may result in mistrust among grant applicants about the review process. Consequently, ensuring transparency is of the utmost importance. Consistency in the review process could also be improved by providing common definitions for each assessment criterion and uniform requirements for grant application submissions. Further research is needed to assess the feasibility and acceptability of these measures.     

Influence of omega-3 Fatty Acid status on the way rats adapt to chronic restraint stress

Omega-3 fatty acids are important for several neuronal and cognitive functions. Altered omega-3 fatty acid status has been implicated in reduced resistance to stress and mood disorders. We therefore evaluated the effects of repeated restraint stress (6 h/day for 21 days) on adult rats fed omega-3 deficient, control or omega-3 enriched diets from conception. We measured body weight, plasma corticosterone and hippocampus glucocorticoid receptors and correlated these data with emotional and depression-like behaviour assessed by their open-field (OF) activity, anxiety in the elevated-plus maze (EPM), the sucrose preference test and the startle response. We also determined their plasma and brain membrane lipid profiles by gas chromatography. Repeated restraint stress caused rats fed a control diet to lose weight. Their plasma corticosterone increased and they showed moderate behavioural changes, with increases only in grooming (OF test) and entries into the open arms (EPM). Rats fed the omega-3 enriched diet had a lower stress-induced weight loss and plasma corticosterone peak, and reduced grooming. Rats chronically lacking omega-3 fatty acid exhibited an increased startle response, a stress-induced decrease in locomotor activity and exaggerated grooming. The brain omega-3 fatty acids increased as the dietary omega-3 fatty acids increased; diets containing preformed long-chain omega-3 fatty acid were better than diets containing the precursor alpha-linolenic acid. However, the restraint stress reduced the amounts of omega-3 incorporated. These data showed that the response to chronic restraint stress was modulated by the omega-3 fatty acid supply, a dietary deficiency was deleterious while enrichment protecting against stress.     

Molecular and Phylogeographic Analysis of Human Immuno-deficiency Virus Type 1 Strains Infecting Treatment-naive Patients from Kigali, Rwanda

This study aimed at describing the genetic subtype distribution of HIV-1 strains circulating in Kigali and their epidemiological link with the HIV-1 strains from the five countries surrounding Rwanda. One hundred and thirty eight pol (RT and PR) sequences from 116 chronically- and 22 recently-infected antiretroviral therapy (ART)-naïve patients from Kigali were generated and subjected to HIV drug resistance (HIV-DR), phylogenetic and recombinant analyses in connection with 366 reference pol sequences from Rwanda, Burundi, Kenya, Democratic Republic of Congo, Tanzania and Uganda (Los Alamos database). Among the Rwandan samples, subtype A1 predominated (71.7%), followed by A1/C recombinants (18.1%), subtype C (5.8%), subtype D (2.9%), one A1/D recombinant (0.7%) and one unknown subtype (0.7%). Thirteen unique and three multiple A1/C recombinant forms were identified. No evidence for direct transmission events was found within the Rwandan strains. Molecular characteristics of HIV-1 were similar between chronically and recently-infected individuals and were not significantly associated with demographic or social factors. Our report suggests that the HIV-1 epidemic in Kigali is characterized by the emergence of A1/C recombinants and is not phylogenetically connected with the HIV-1 epidemic in the five neighboring countries. The relatively low level of transmitted HIV-DR mutations (2.9%) reported here indicates the good performance of the ART programme in Rwanda. However, the importance of promoting couples'' counseling, testing and disclosure during HIV prevention strategies is highlighted.     

HIV Protease Inhibitors Do Not Cause the Accumulation of Prelamin A in PBMCs from Patients Receiving First Line Therapy: The ANRS EP45 “Aging” Study

Background

The ANRS EP45 “Aging” study investigates the cellular mechanisms involved in the accelerated aging of HIV-1 infected and treated patients. The present report focuses on lamin A processing, a pathway known to be altered in systemic genetic progeroid syndromes.

Methods

35 HIV-1 infected patients being treated with first line antiretroviral therapy (ART, mean duration at inclusion: 2.7±1.3 years) containing boosted protease inhibitors (PI/r) (comprising lopinavir/ritonavir in 65% of patients) were recruited together with 49 seronegative age- and sex-matched control subjects (http://clinicaltrials.gov/, {"type":"clinical-trial","attrs":{"text":"NCT01038999","term_id":"NCT01038999"}}NCT01038999). In more than 88% of patients, the viral load was <40 copies/ml and the CD4+ cell count was >500/mm³. Prelamin A processing in peripheral blood mononuclear cells (PBMCs) from patients and controls was analysed by western blotting at inclusion. PBMCs from patients were also investigated at 12 and 24 months after enrolment in the study. PBMCs from healthy controls were also incubated with boosted lopinavir in culture medium containing various concentrations of proteins (4 to 80 g/L).

Results

Lamin A precursor was not observed in cohort patient PBMC regardless of the PI/r used, the dose and the plasma concentration. Prelamin A was detected in PBMC incubated in culture medium containing a low protein concentration (4 g/L) but not in plasma (60–80 g/L) or in medium supplemented with BSA (40 g/L), both of which contain a high protein concentration.

Conclusions

Prelamin A processing abnormalities were not observed in PBMCs from patients under the PI/r first line regimen. Therefore, PI/r do not appear to contribute to lamin A-related aging in PBMCs. In cultured PBMCs from healthy donors, prelamin A processing abnormalities were only observed when the protein concentration in the culture medium was low, thus increasing the amount of PI available to enter cells.ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01038999","term_id":"NCT01038999"}}NCT01038999 http://clinicaltrials.gov/ct2/show/{"type":"clinical-trial","attrs":{"text":"NCT01038999","term_id":"NCT01038999"}}NCT01038999.     

Bivalirudin in combination with heparin to control mesenchymal cell procoagulant activity

Islet and hepatocyte transplantation are associated with tissue factor-dependent activation of coagulation which elicits instant blood mediated inflammatory reaction, thereby contributing to a low rate of engraftment. The aim of this study was i) to evaluate the procoagulant activity of human adult liver-derived mesenchymal progenitor cells (hALPCs), ii) to compare it to other mesenchymal cells of extra-hepatic (bone marrow mesenchymal stem cells and skin fibroblasts) or liver origin (liver myofibroblasts), and iii) to determine the ways this activity could be modulated. Using a whole blood coagulation test (thromboelastometry), we demonstrated that all analyzed cell types exhibit procoagulant activity. The hALPCs pronounced procoagulant activity was associated with an increased tissue factor and a decreased tissue factor pathway inhibitor expression as compared with hepatocytes. At therapeutic doses, the procoagulant effect of hALPCs was inhibited by neither antithrombin activators nor direct factor Xa inhibitor or direct thrombin inhibitors individually. However, concomitant administration of an antithrombin activator or direct factor Xa inhibitor and direct thrombin inhibitor proved to be a particularly effective combination for controlling the procoagulant effects of hALPCs both in vitro and in vivo. The results suggest that this dual antithrombotic therapy should also improve the efficacy of cell transplantation in humans.     

In vivo importance of homologous recombination DNA repair for mouse neural stem and progenitor cells

We characterized the in vivo importance of the homologous recombination factor RAD54 for the developing mouse brain cortex in normal conditions or after ionizing radiation exposure. Contrary to numerous homologous recombination genes, Rad54 disruption did not impact the cortical development without exogenous stress, but it dramatically enhanced the radiation sensitivity of neural stem and progenitor cells. This resulted in the death of all cells irradiated during S or G2, whereas the viability of cells irradiated in G1 or G0 was not affected by Rad54 disruption. Apoptosis occurred after long arrests at intra-S and G2/M checkpoints. This concerned every type of neural stem and progenitor cells, showing that the importance of Rad54 for radiation response was linked to the cell cycle phase at the time of irradiation and not to the differentiation state. In the developing brain, RAD54-dependent homologous recombination appeared absolutely required for the repair of damages induced by ionizing radiation during S and G2 phases, but not for the repair of endogenous damages in normal conditions. Altogether our data support the existence of RAD54-dependent and -independent homologous recombination pathways.