Serge Milin

Prognostic value of increase in transcript levels of Tp73 ΔEx2-3 isoforms in low-grade glioma patients

Glial tumours are a devastating, poorly understood condition carrying a gloomy prognosis for which clinicians sorely lack reliable predictive parameters facilitating a sound treatment strategy. Tp73, a p53 family member, expresses two main classes of isoforms – transactivatory activity (TA)p73 and ΔTAp73 – exhibiting tumour suppressor gene and oncogene properties, respectively. The authors examined their expression status in high- and low-grade adult gliomas. Isoform-specific real-time reverse transcription-polymerase chain reaction was used for the analysis of Tp73 isoform transcript expression in a series of 51 adult patients harbouring glial tumours, in order to compare tumour grades with each other, and with non-tumoural samples obtained from epileptic patients as well. Our data demonstrate increase of TAp73 and ΔTAp73 transcript levels at onset and early stage of the disease. We also show that ΔEx2–3 isoform expression in low-grade tumours anticipates clinical and imaging progression to higher grades, and correlates to the patients’ survival. Expression levels of P1 promoter generated Tp73 isoforms – and particularly ΔEx2–3 – indeed allow for prediction of the clinical progression of low-grade gliomas in adults. Our data are the first such molecular biology report regarding low-grade tumours and as such should be of help for sound decision-making.

Direct Thrombin Inhibitor Prevents Delayed Graft Function in a Porcine Model of Renal Transplantation

Background. Kidney transplantations from donors after cardiac arrest (DCA) are characterized by an increase in the occurrence of delayed graft function and primary nonfunction. In this study, Melagatran, a selective reversible direct thrombin inhibitor was used to limit renal injury in a DCA pig kidney transplantation model. Methods. We used a porcine model of DCA to study the effects of treatment with Melagatran in the peri-conservation period. Thromboelastography was used to check Melagatran antithrombin effect on in vitro clot formation. Reverse-transcriptase polymerase chain reaction was used to analyze the peripheral immune cells activation status. Renal function and morphologic study were performed at days 1 and 7. Finally, we analyzed the mechanisms of Melagatran protection on kidney microvasculature primary endothelial cells. Results. Prolongation of coagulation time (Ex-Tem) was observed 10 min after injection; however, Melagatran did not modulate increases of thrombin-antithrombin complexes following reperfusion. Melagatran significant treatment lowered the proinflammatory status of circulating immune cells. Animals survival was increased in Melagatran-treated groups (9 of 10 in Melagatran groups vs. 4 of 10 in controls at day 7). Renal injury and inflammation were also significantly reduced in treated groups. We also demonstrated a direct protective effect of Melagatran against endothelial cell activation and inflammation in vitro. Conclusion. Direct thrombin inhibitor administration in the periconservation period improved graft outcome and reduced renal injury in a model of DCA.

Prognostic molecular markers with no impact on decision-making: the paradox of gliomas based on a prospective study

This study assessed the prognostic value of several markers involved in gliomagenesis, and compared it with that of other clinical and imaging markers already used. Four-hundred and sixteen adult patients with newly diagnosed glioma were included over a 3-year period and tumour suppressor genes, oncogenes, MGMT and hTERT expressions, losses of heterozygosity, as well as relevant clinical and imaging information were recorded. This prospective study was based on all adult gliomas. Analyses were performed on patient groups selected according to World Health Organization histoprognostic criteria and on the entire cohort. The endpoint was overall survival, estimated by the Kaplan–Meier method. Univariate analysis was followed by multivariate analysis according to a Cox model. p14ARF, p16INK4A and PTEN expressions, and 10p 10q23, 10q26 and 13q LOH for the entire cohort, hTERT expression for high-grade tumours, EGFR for glioblastomas, 10q26 LOH for grade III tumours and anaplastic oligodendrogliomas were found to be correlated with overall survival on univariate analysis and age and grade on multivariate analysis only. This study confirms the prognostic value of several markers. However, the scattering of the values explained by tumour heterogeneity prevents their use in individual decision-making.

Analysis of machine perfusion benefits in kidney grafts: a preclinical study

BackgroundMachine perfusion (MP) has potential benefits for marginal organs such as from deceased from cardiac death donors (DCD). However, there is still no consensus on MP benefits. We aimed to determine machine perfusion benefits on kidney grafts.MethodsWe evaluated kidney grafts preserved in ViaspanUW or KPS solutions either by CS or MP, in a DCD pig model (60 min warm ischemia + 24 h hypothermic preservation). Endpoints were: function recovery, quality of function during follow up (3 month), inflammation, fibrosis, animal survival.ResultsViaspanUW-CS animals did not recover function, while in other groups early follow up showed similar values for kidney function. Alanine peptidase and β-NAG activities in the urine were higher in CS than in MP groups. Oxydative stress was lower in KPS-MP animals. Histology was improved by MP over CS. Survival was 0% in ViaspanUW-CS and 60% in other groups. Chronic inflammation, epithelial-to-mesenchymal transition and fibrosis were lowest in KPS-MP, followed by KPS-CS and ViaspanUW-MP.ConclusionsWith ViaspanUW, effects of MP are obvious as only MP kidney recovered function and allowed survival. With KPS, the benefits of MP over CS are not directly obvious in the early follow up period and only histological analysis, urinary tubular enzymes and red/ox status was discriminating. Chronic follow-up was more conclusive, with a clear superiority of MP over CS, independently of the solution used. KPS was proven superior to ViaspanUW in each preservation method in terms of function and outcome. In our pre-clinical animal model of DCD transplantation, MP offers critical benefits.

A mesenchymal glioma stem cell profile is related to clinical outcome

Recent studies have demonstrated a relationship between the expression of stem cell-associated genes and relapses in glioblastoma (GBM), suggesting a key role for tumor stem cells in this process. Although there is increasing interest in this field, glioma stem cells (GSCs) are still poorly characterized, their ‘stemness’ state and factors maintaining these properties remain largely unknown. We performed an expression profiling analysis of pluripotency in gliomaspheres derived from 11 patients. Comparative analysis between GSCs and H1 and H9 human embryonic stem cells as well as H9-derived neural stem cells indicates major variations in gene expression of pluripotency factors Nanog and OCT4, but a stable pattern for SOX2 suggesting its important function in maintaining pluripotency in GSCs. Our results also showed that all GSC lines have the capacity to commit to neural differentiation and express mesenchymal or endothelial differentiation markers. In addition, hierarchical clustering analysis revealed two groups of GSCs reflecting their heterogeneity and identified COL1A1 and IFITM1 as the most discriminating genes. Similar patterns have been observed in tumors from which gliomaspheres have been established. To determine whether this heterogeneity could be clinically relevant, the expression of both genes was further analyzed in an independent cohort of 30 patients with GBM and revealed strong correlation with overall survival. In vitro silencing of COL1A1 and IFTM1 confirmed the effect of these mesenchymal-associated genes on cell invasion and gliomasphere initiation. Our results indicate that COL1A1 and IFITM1 genes could be considered for use in stratifying patients with GBM into subgroups for risk of recurrence at diagnosis, as well as for prognostic and therapeutic evolution.

A p38 mitogen-activated protein kinase inhibitor protects against renal damage in a non-heart-beating donor model

Ischemia-reperfusion injury is one of the central nonimmunologic processes involved in renal allograft dysfunction. Kidneys from non-heart beating donors (NHBD) exhibit higher rates of delayed graft function (DGF) than those from other donors. Primary nonfunction and DGF are the main barriers to the use of kidneys from NHBD. Using a pig model of NHBD transplantation, we studied the effect of FR167653 (a p38 MAP kinase inhibitor) on the recovery and reparation of kidneys exposed to both warm (WI: 1 h) and cold ischemia (24 h). Our results demonstrate that the addition of FR167653 increases the kinetics of proximal tubule cell regeneration after 60 min of WI. Hypoxia-inducible factor and vascular endothelial growth factor expression was also more important in FR167653-treated kidneys compared with those in nontreated groups. Also, expression of peripheral-type benzodiazepine receptor, involved in tissue repair, was increased in the FR167653-treated groups. At 3 mo, the protective effects of FR167653 were accompanied by a reduction of long-term inflammation process and tubulointerstitial fibrosis development associated with a limitation of ischemia-induced remodeling. This study suggests that such treatment may be useful in protocols aimed at improving the quality of renal transplants from NHBD. In addition, the beneficial role of FR167653 in limiting early injury is associated with secondary reduction in development of tubular atrophy and interstitial fibrosis which are together the hallmark of failing renal transplants. The more efficient effect was observed when FR167653 was added in combination before WI, during cold storage and reperfusion.

Supratentorial clear cell ependymomas with branching capillaries demonstrate characteristic clinicopathological features and pathological activation of nuclear factor-kappaB signaling

BACKGROUND Clear cell ependymoma is one of the 4 main histological subtypes of ependymomas defined by the World Health Organization (WHO) classification of tumors of the CNS. DNA methylation profiling can distinguish 4 subgroups of intracranial ependymomas, including supratentorial (ST) ependymomas with Yes-associated protein 1 fusion (YAP1), ST ependymomas with fusion of v-rel avian reticuloendotheliosis viral oncogene homolog A (RELA), posterior fossa ependymomas with balanced genome, and posterior fossa ependymomas with chromosomal instability. In addition, trisomy 19 is a genomic hallmark of ependymomas with rich branching capillaries. However, the relation of histological and molecular subtypes is unclear. METHODS Here, we report a series of 20 ependymomas histologically defined by clear cells and branching capillaries. RESULTS We observed a strong male predominance. Median age at surgery was 10.4 years (range, 0.8-68.4). All cases were ST, cortical, contrast enhancing, and most often frontal, cystic, and calcified. All tumors qualified as WHO grade III. Some of them exhibited neuronal differentiation. Trisomy 19 was recorded in 13 cases. All samples strongly accumulated p65RelA protein within nuclei, indicating pathological activation of the nuclear factor-kappaB pathway. We identified causative C11ORF95-RELA fusion in almost all cases. Median progression-free survival and overall survival were 11.4 years (95% CI: 5.1-17.8) and not reached, respectively. CONCLUSION ST clear cell ependymomas with branching capillaries display characteristic clinicopathological features and are associated with pathological activation of nuclear factor-kappaB signaling, which may indicate a potential novel target for therapy in these patients.

The use of impression cytology in the follow‐up of severe ocular burns

Aims: To evaluate by impression cytology (IC) the expression of the MHC class II inflammatory marker HLA-DR by the conjunctival epithelium, the cytological modifications of the conjunctival surface according to the Nelson’s classification, and the eventual correlation between the two after severe ocular burns. Methods: A total of 24 patients (24 eyes) who presented with severe ocular burns underwent IC. We compared them with 18 healthy eyes. HLA-DR expression was studied by flow cytometry as well as the conjunctival histology evaluated with the Nelson’s classification from 2–24 months after the onset of burns. Results: There was a significant upregulation of the expression of HLA-DR in eyes with burns compared to the healthy population at 2 months (p<0.001), 6 months (p<0.001), 12 months (p = 0.019), 18 months (p = 0.0171) and 24 months (p = 0.01766). A significant difference was found between the Nelson grade in the pathological population and those of the healthy population at 2 months (p = 0.0157). HLA-DR upregulation was significantly correlated with the Nelson’s grades between 2 months (r = 0.69, p<0.0001) and 6 months (r = 0.61, p = 0.0001). Conclusion: The IC technique can act as a useful tool for following-up ocular surface inflammation after severe ocular burns.

Selective Release of a Cyclopamine Glucuronide Prodrug toward Stem-like Cancer Cell Inhibition in Glioblastoma

Recent data suggest that inhibition of the Hedgehog pathway could be a therapeutic target for glioblastoma. Alkaloid cyclopamine inhibits Hedgehog signaling, depleting stem-like cancer cells derived from glioblastoma. However, this compound is toxic for somatic stem cells, preventing its use for clinical applications. In this study, we tested a derivatization product of cyclopamine in the form of cyclopamine glucuronide prodrug (CGP-2). This compound was used in vitro and in vivo toward glioblastoma-initiating cells (GIC). Results obtained in vitro indicate that CGP-2 is active only in the presence of β-glucuronidase, an enzyme detected in high levels in necrotic areas of glioblastomas. CGP-2 decreased proliferation and inhibited the self-renewal of all GIC lines tested. Hedgehog pathway blockade by 10 μmol/L of CGP-2 induced a 99% inhibition of clonogenicity on GICs, similar to cyclopamine treatment. Combination of CGP-2 with radiation decreased clonogenic survival in all GIC lines compared with CGP-2 alone. In a subcutaneous glioblastoma xenograft model, a two-week CGP-2 treatment prevented tumor growth with 75% inhibition at 8 weeks, and this inhibition was still significant after 14 weeks. Unlike cyclopamine, CGP-2 had no detectable toxic effects in intestinal crypts. Our study suggests that inhibition of the Hedgehog pathway with CGP-2 is more effective than conventional temozolomide adjuvant, with much lower concentrations, and seems to be an effective therapeutic strategy for targeting GICs. Mol Cancer Ther; 13(9); 2159–69. ©2014 AACR.

Pleural lipoma: a non-surgical lesion?

Pleural lipomas are benign tumours that develop at the expense of adipose tissues, and they never evolve towards liposarcoma. Located usually at the mediastinal, bronchial and pulmonary levels, a pleural situation is extremely rare. Chest X-rays usually detect them and computed tomography scans confirm the diagnosis. As complications occur, a wait-and-see policy is common. We report our pleural lipoma surgical exeresis experience since 1999. We have operated on five cases of pleural lipomas among nearly 1800 cases of thoracic exeresis: three male and two female patients, without obesity (in all cases, body mass index (BMI) < 28). The mean age was 54.6 years (range 35-72 years). Four patients were electively operated and one in emergency, three with video-assisted thoracic surgery (VATS) procedure and two with open chest surgery, without recurrent cases. Advancements in VATS have greatly reduced the morbidity rate of these benign tumours especially if exeresis is performed early on a small, uncomplicated adhesion-free tumour. On the other hand, the operation may be deleterious, complicated by the presence of a large lipoma or in a complicating situation. In our opinion, we should revise the wait-and-see policy when facing these lesions considering their evolutionary potential. We should advise VATS in pleural lipomas.