Hugues Loiseau

Experimental anti-angiogenesis causes upregulation of genes associated with poor survival in glioblastoma

Vascular endothelial growth factor (VEGF) inhibitors are the most promising anti‐angiogenic agents used increasingly in the clinic. However, to be efficient, anti‐VEGF agents need to be associated with classic chemotherapy. Exploring gene regulation in tumor cells during anti‐angiogenesis might help to comprehend the molecular basis of response to treatment. To generate a defined anti‐angiogenic condition in vivo, we transfected human glioma cells with short‐interfering RNAs against VEGF‐A and implanted them on the chick chorio‐allantoic membrane. Gene regulation in avascular tumors was studied using human Affymetrix™ GeneChips. Potentially important genes were further studied in glioma patients. Despite strong VEGF inhibition, we observed recurrent formation of small, avascular tumors. CHI3L2, IL1B, PI3/elafin and CHI3L1, which encodes for YKL‐40, a putative prognosticator for various diseases, including cancer, were strongly up‐regulated in avascular glioma. In glioblastoma patients, these genes showed coregulation and their expression differed significantly from low‐grade glioma. Importantly, high levels of CHI3L1 (p = 0.036) and PI3/elafin mRNA (p = 0.0004) were significantly correlated with poor survival. Cox regression analysis further confirmed that PI3 and CHI3L1 levels are survival markers independent from patient age and sex. Elafin‐positive tumor cells were only found in glioblastoma, where they were clustered around necrotic areas. PI3/elafin is strongly induced by serum deprivation and hypoxia in U87 glioma cells in vitro. Our results indicate that anti‐angiogenesis in experimental glioma drives expression of critical genes which relate to disease aggressiveness in glioblastoma patients. In particular, CHI3L1 and PI3/elafin may be useful as new prognostic markers and new therapeutic targets.

Mobile phone use and brain tumours in the CERENAT case-control study

The carcinogenic effect of radiofrequency electromagnetic fields in humans remains controversial. However, it has been suggested that they could be involved in the aetiology of some types of brain tumours. Objectives The objective was to analyse the association between mobile phone exposure and primary central nervous system tumours (gliomas and meningiomas) in adults. Methods CERENAT is a multicenter case-control study carried out in four areas in France in 2004–2006. Data about mobile phone use were collected through a detailed questionnaire delivered in a face-to-face manner. Conditional logistic regression for matched sets was used to estimate adjusted ORs and 95% CIs. Results A total of 253 gliomas, 194 meningiomas and 892 matched controls selected from the local electoral rolls were analysed. No association with brain tumours was observed when comparing regular mobile phone users with non-users (OR=1.24; 95% CI 0.86 to 1.77 for gliomas, OR=0.90; 95% CI 0.61 to 1.34 for meningiomas). However, the positive association was statistically significant in the heaviest users when considering life-long cumulative duration (≥896u2005h, OR=2.89; 95% CI 1.41 to 5.93 for gliomas; OR=2.57; 95% CI 1.02 to 6.44 for meningiomas) and number of calls for gliomas (≥18u2005360 calls, OR=2.10, 95% CI 1.03 to 4.31). Risks were higher for gliomas, temporal tumours, occupational and urban mobile phone use. Conclusions These additional data support previous findings concerning a possible association between heavy mobile phone use and brain tumours.

Brain tumours and exposure to pesticides: a case-control study in southwestern France.

Background: Brain tumours are often disabling and rapidly lethal; their aetiology is largely unknown. Among potential risk factors, pesticides are suspected. Objective: To examine the relationship between exposure to pesticides and brain tumours in adults in a population-based case–control study in southwestern France. Methods: Between May 1999 and April 2001, 221 incident cases of brain tumours and 442 individually matched controls selected from the general population were enrolled. Histories of occupational and environmental exposures, medical and lifestyle information were collected. A cumulative index of occupational exposure to pesticides was created, based on expert review of lifelong jobs and tasks. Separate analyses were performed for gliomas and meningiomas. Results: A non-statistically significant increase in risk was found for brain tumours when all types of occupational exposure to pesticides were considered (ORu200a=u200a1.29, 95% CI 0.87 to 1.91) and slightly higher but still non-statistically significant when gliomas were considered separately (ORu200a=u200a1.47, 95% CI 0.81 to 2.66). In the highest quartile of the cumulative index, a significant association was found for brain tumours (ORu200a=u200a2.16, 95% CI 1.10 to 4.23) and for gliomas (ORu200a=u200a3.21, 95% CI 1.13 to 9.11), but not for meningiomas. A significant increase in risk was also seen for the treatment of home plants (ORu200a=u200a2.24, 95% CI 1.16 to 4.30) owing to environmental exposure to pesticides. Conclusions: These data suggest that a high level of occupational exposure to pesticides might be associated with an excess risk of brain tumours, and especially of gliomas.

Incidence of central nervous system tumors in Gironde, France

The incidence of CNS tumors is subject to geographical and temporal variations which are poorly understood. The incidence of these tumors was studied in Gironde, a department of southwestern France with 1,058,911 inhabitants older than 16 years. We recorded any malignant or benign central nervous system (CNS) tumor diagnosed between May 1999 and April 2001 in adults living in Gironde. Three hundred and twenty-nine CNS tumors were diagnosed during the study period. The incidence of CNS tumors in adults was therefore 15.5 per 100,000. Overall, the incidence according to sex was 14.7 for males and 16.2 per 100,000 for females. The incidence rate increased according to age up to 80 years (3.7 per 100,000 for 20–29 years of age to 33.4 per 100,000 for 70–79 years of age) and reduced thereafter for gliomas. Our estimation of the incidence of CNS tumors ranks high among the earlier reports. Further analytic studies are ongoing.

Brain tumors and hormonal factors: review of the epidemiological literature

BackgroundTo date, the etiology of primary tumors of the central nervous system (mainly gliomas and meningiomas) is poorly understood. The role of sex hormones has been suggested, based on clinical, experimental, biological, and epidemiological data.ObjectiveTo review the epidemiological studies on the relation between hormonal factors and the occurrence of glioma and meningioma, in order to identify new research developments.MethodsArticles published until September 2010 were selected by considering exogenous and endogenous exposures and specific brain tumors. Standardized information was collected from 20 articles: 15 concerning gliomas and 13 meningiomas.ResultsAn increased glioma risk was observed with later menarche and menopause, while a reduced glioma risk was observed with hormone replacement therapy (HRT) and oral contraceptive use, despite duration of use had no effect on risk. Meningioma risk increased after menopause and with HRT use. No clear association was found with pregnancy and breastfeeding.ConclusionResults are globally concordant with the biologic hypothesis assuming that female sex hormones are protective against glioma and may increase the risk of meningioma. However, new epidemiological studies should be conducted in order to confirm these associations and to refine the role of hormonal factors in brain etiology.

FBXW7/hCDC4 controls glioma cell proliferation in vitro and is a prognostic marker for survival in glioblastoma patients

BackgroundIn the quest for novel molecular mediators of glioma progression, we studied the regulation of FBXW 7 (hCDC 4/hAGO/SEL 10), its association with survival of patients with glioblastoma and its potential role as a tumor suppressor gene in glioma cells. The F-box protein Fbxw7 is a component of SCFFbxw7, a Skp1-Cul1-F-box E3 ubiquitin ligase complex that tags specific proteins for proteasome degradation. FBXW 7 is mutated in several human cancers and functions as a haploinsufficient tumor suppressor in mice. Any of the identified targets, Cyclin E, c-Myc, c-Jun, Notch1/4 and Aurora-A may have oncogenic properties when accumulated in tumors with FBXW 7 loss.ResultsWe tested the expression of FBXW 7 in human glioma biopsies by quantitative PCR and compared the transcript levels of grade IV glioma (glioblastoma, G-IV) with those of grade II tumors (G-II). In more than 80% G-IV, expression of FBXW 7 was significantly reduced. In addition, levels of FBXW 7 were correlated with survival indicating a possible implication in tumor aggressiveness. Locus 4q31.3 which carries FBXW 7 was investigated by in situ hybridization on biopsy touchprints. This excluded allelic loss as the principal cause for low expression of FBXW 7 in G-IV tumors. Two targets of Fbxw7, Aurora-A and Notch4 were preferentially immunodetected in G-IV biopsies. Next, we investigated the effects of FBXW 7 misregulation in glioma cells. U87 cells overexpressing nuclear isoforms of Fbxw7 lose the expression of the proliferation markers PCNA and Ki-67, and get counterselected in vitro. This observation fits well with the hypothesis that Fbxw7 functions as a tumor suppressor in astroglial cells. Finally, FBXW 7 knockdown in U87 cells leads to defects in mitosis that may promote aneuploidy in progressing glioma.ConclusionOur results show that FBXW 7 expression is a prognostic marker for patients with glioblastoma. We suggest that loss of FBXW 7 plays an important role in glioma malignancy by allowing the accumulation of multiple oncoproteins and that interfering with Fbxw7 or its downstream targets would constitute a new therapeutic advance.

Autophagy inhibition cooperates with erlotinib to induce glioblastoma cell death

Gliomas are the most common malignant primary brain tumors in adults. The median survival never exceeds 12 months, owing to inherent resistance to both radio and chemotherapies. Epidermal Growth Factor Receptor (EGFR) is amplified, overexpressed, and/or mutated in glioblastomas (GBM), making it a rational for therapy. Erlotinib, an EGFR kinase inhibitor is strongly associated with clinical response in several cancers. Inhibition of cell proliferation and induction of apoptosis by erlotinib were investigated in U87-MG and DBTRG-05MG, two human glioblastoma cell lines. The expression of several apoptosis-related proteins was investigated in these cell lines and in tumoral tissue from glioblastomas. Both cell lines expressed wild-type EGFR but were deficient for PTEN. Erlotinib induced a marked accumulation of the BIM protein, but the activation of caspase-3 machinery was missing, regardless of the decrease in XIAP. Moreover, in U87-MG, erlotinib promoted accumulation of αB-crystallin a small heat shock protein capable to impair caspase activation. DBTRG-05MG was found deficient for procaspase 3 and constitutively overexpressed αB-crystallin. Similarly, deficiencies in PTEN and procaspase 3 were constantly found in samples from glioblastoma samples, while αB-crystallin expression was inconsistent. In cell lines, high concentrations of erlotinib induced cell death through a caspase independent process and an autophagic process was evidenced in U87-MG. Inhibition of autophagy induced a marked increase in the death-inducing activity of erlotinib. These results confirm that glioblastoma cell lines exhibit several anti-apoptotic mechanisms, and underline that EGFR targeted therapy must be associated to other inhibitors to achieve an antitumoral effect.

Occupational and residential exposure to electromagnetic fields and risk of brain tumors in adults: a case-control study in Gironde, France

The etiology of brain tumors remains largely unknown. Among potential risk factors, exposure to electromagnetic fields is suspected. We analyzed the relationship between residential and occupational exposure to electromagnetic field and brain tumors in adults. A case–control study was carried out in southwestern France between May 1999 and April 2001. A total of 221 central nervous system tumors (105 gliomas, 67 meningiomas, 33 neurinomas and 16 others) and 442 individually age‐ and sex‐matched controls selected from general population were included. Electromagnetic field exposure [extremely low frequency (ELF) and radiofrequency separately was assessed in occupational settings through expert judgement based on complete job calendar, and at home by assessing the distance to power lines with the help of a geographical information system. Confounders such as education, use of home pesticide, residency in a rural area and occupational exposure to chemicals were taken into account. Separate analyses were performed for gliomas, meningiomas and acoustic neurinomas. A nonsignificant increase in risk was found for occupational exposure to electromagnetic fields [odds ratio (OR = 1.52, 0.92–2.51)]. This increase became significant for meningiomas, especially when considering ELF separately [OR = 3.02; 95 percent confidence interval (95% CI) =1.10–8.25]. The risk of meningioma was also higher in subjects living in the vicinity of power lines (<100 m), even if not significant (OR = 2.99, 95% CI 0.86–10.40). These data suggest that occupational or residential exposure to ELF may play a role in the occurrence of meningioma.

Oncological patterns of care and outcomes for 265 elderly patients with newly diagnosed glioblastoma in France

The incidence of glioblastoma (GBM) has increased in patients aged 70xa0years or older, and will continue to grow. Elderly GBM patients have been excluded from most clinical trials; furthermore, optimal care management as well as benefit/risk ratio of GBM treatments are still being debated. This study describes oncological patterns of care, prognostic factors, and survival for patients ≥70xa0years in France. We identified patients over 70 with newly diagnosed and histologically confirmed GBM on data previously published by the French Brain Tumor DataBase. We included 265 patients. Neurological deficits and mental status disorders were the most frequent symptoms. The surgery consisted of resection (RSnu2009=u200995) or biopsy (Bnu2009=u2009170); 98 patients did not have subsequent oncological treatment. After surgery, first-line treatment consisted of radiotherapy (RTnu2009=u200976), chemotherapy (CTnu2009=u200952), and concomitant radiochemotherapy (CRCnu2009=u200939). The median age at diagnosis was 76, 74, and 73xa0years, respectively, for the untreated, Bu2009+u2009RT and/or CT, RSu2009±u2009RT and/or CT groups. Median survival (in days, 95xa0% CI) with these main strategies, when analyzed according to surgical groups, was: B-CTnu2009=u200941, 199[155–280]; B-CRCnu2009=u200921, 318[166–480]; B-RTnu2009=u200937, 149[130–214]; RS-CTnu2009=u200911, 245[211–na]; RS-CRCnu2009=u200918, 372[349–593]; RS-RTnu2009=u200939, 269[218–343]. This population study for elderly GBM patients is one of the most important in Europe, and could be considered as a historical cohort to compare future treatments. Moreover, we can hypothesize that elderly patients (versus patients <70xa0years) are undertreated. Karnofsky performance status seems to be the most relevant clinical predictive factor, and RS and CRC have a positive impact on survival for elderly GBM patients in the general population, at least when feasible.

99mTc-MAG3-Aptamer for Imaging Human Tumors Associated with High Level of Matrix Metalloprotease-9

The human matrix metalloprotease 9 (hMMP-9) is involved in many physiological processes such as tissue remodeling. Its overexpression in tumors promotes the release of cancer cells thus contributing to tumor metastasis. It is a relevant marker of malignant tumors. We selected an RNA aptamer containing 2-fluoro, pyrimidine ribonucleosides, that exhibits a strong affinity for hMMP-9 (K(d) = 20 nM) and that discriminates other human MMPs: no binding was detected to either hMMP-2 or -7. Investigating the binding properties of different MMP-9 aptamer variants by surface plasmon resonance allowed the determination of recognition elements. As a result, a truncated aptamer, 36 nucleotides long, was made fully resistant to nuclease following the substitution of every purine ribonucleoside residue by 2-O-methyl analogues and was conjugated to S-acetylmercaptoacetyltriglycine for imaging purposes. The resulting modified aptamer retained the binding properties of the originally selected sequence. Following (99m)Tc labeling, this aptamer was used for ex vivo imaging slices of human brain tumors. We were able to specifically detect the presence of hMMP-9 in such tissues.