Navigating the highlights of phase III trials: a watchful eye on evidence-based radiotherapy.

BACKGROUND: Phase III randomized controlled trials (RCTs) are the cornerstone of evidence-based oncology. However, there is no exhaustive review describing the radiotherapy RTCs characteristics. The objective of the present study was to describe features of all phase III RCTs including at least a radiation therapy. METHODS AND MATERIALS: Requests were performed in the Medline database (via PubMed). The latest update was performed in April 2016, using the following MESH terms: 'clinical trials: phase III as topic', 'radiotherapy', 'brachytherapy', as keywords. RESULTS: A total of 454 phase III RCTs were identified. Studies were mainly based on open (92.1%) multicenter (77.5%) designs, analyzed in intend to treat (67.6%), aiming at proving superiority (91.6%) through overall survival assessment (46.5%). Most frequently studied malignancies were head and neck (21.8%), lung (14.3%) and prostate cancers (9.9%). Patients were mainly recruited with a locally advanced disease (73.7%). Median age was 59 years old. Out of 977 treatment arms, 889 arms experienced radiotherapy, mainly using 3D-conformal radiotherapy (288 arms, 32.4%). Intensity-modulated techniques were tested in 12 arms (1.3%). The intervention was a non-cytotoxic agent addition in 89 studies (19.6%), a radiation dose/fractionation modification in 74 studies (16.3%), a modification of chemotherapy regimen in 63 studies (13.9%), a chemotherapy addition in 63 studies (13.9%) and a radiotherapy addition in 53 trials (11.7%). With a median follow-up of 50 months, acute all-grade and grade 3-5 toxicities were reported in 49.6% and 69.4% of studies, respectively. Radiotherapy technique, follow-up and late toxicities were reported in 60.1%, 74%, and 31.1% of studies, respectively. CONCLUSION: Phase III randomized controlled trials featured severe limitations, since a third did not report radiotherapy technique, follow-up or late toxicities. The fast-paced technological evolution creates a discrepancy between literature and radiotherapy techniques performed in daily-routine, suggesting that phase III methodology needs to be reinvented.

In vitro radiosensitizing effects of ultrasmall gadolinium based particles on tumour cells.

Since radiotherapy is widely used in cancer treatment, it is essential to develop strategies which lower the irradiation burden while increasing efficacy and become efficient even in radio resistant tumors. Our new strategy is relying on the development of solid hybrid nanoparticles based on rare-earth such as gadolinium. In this paper, we then evidenced that gadolinium-based particles can be designed to enter efficiently into the human glioblastoma cell line U87 in quantities that can be tuned by modifying the incubation conditions. These sub-5 nm particles consist in a core of gadolinium oxide, a shell of polysiloxane and are functionalized by diethylenetriaminepentaacetic acid (DTPA). Although photoelectric effect is maximal in the [10-100 keV] range, such particles were found to possess efficient in-vitro radiosensitizing properties at an energy of 660 keV by using the "single-cell gel electrophoresis comet assay," an assay that measures the number of DNA damage that occurs during irradiation. Even more interesting, the particles have been evidenced by MTT assays to be also efficient radiosensitizers at an energy of 6 MeV for doses comprised between 2 and 8 Gy. The properties of the gadolinium-based particles give promising opening to a particle-assisted radio-therapy by using irradiation systems already installed in the majority of hospitals.

High expression of gabarapl1 is associated with a better outcome for patients with lymph node-positive breast cancer.

BACKGROUND: This study evaluates the relation of the early oestrogen-regulated gene gabarapl1 to cellular growth and its prognostic significance in breast adenocarcinoma. METHODS: First, the relation between GABARAPL1 expression and MCF-7 growth rate was analysed. Thereafter, by performing macroarray and reverse transcriptase quantitative-polymerase chain reaction (RT-qPCR) experiments, gabarapl1 expression was quantified in several histological breast tumour types and in a retrospective cohort of 265 breast cancers. RESULTS: GABARAPL1 overexpression inhibited MCF-7 growth rate and gabarapl1 expression was downregulated in breast tumours. Gabarapl1 mRNA levels were found to be significantly lower in tumours presenting a high histological grade, with a lymph node-positive (pN+) and oestrogen and/or progesterone receptor-negative status. In univariate analysis, high gabarapl1 levels were associated with a lower risk of metastasis in all patients (hazard ratio (HR) 4.96), as well as in pN+ patients (HR 14.96). In multivariate analysis, gabarapl1 expression remained significant in all patients (HR 3.63), as well as in pN+ patients (HR 5.65). In univariate or multivariate analysis, gabarapl1 expression did not disclose any difference in metastasis risk in lymph node-negative patients. CONCLUSIONS: Our data show for the first time that the level of gabarapl1 mRNA expression in breast tumours is a good indicator of the risk of recurrence, specifically in pN+ patients.

Gadolinium-Based Nanoparticles Can Overcome the Radioresistance of Head and Neck Squamous Cell Carcinoma Through the Induction of Autophagy.

Radiation therapy is a mainstay in the therapeutic management of Head and Neck Squamous Cell Carcinoma (HNSCC). Despite significant progress in this field, radioresistance still accounts for most treatment failures. Gadolinium-based nanoparticles (GBNs) have shown great promises as radiosensitizers but the underlying sensitizing mechanism is still largely unknown with regards to the disparities obtained in in vitro studies. In this study, we show that a new formulation of GBNs, AGuIX®, can radiosensitize HNSCC after cell uptake and further accumulation in lysosomes. Although radiation alone triggered late apoptosis and mitochondrial impairment, the pre-treatment with GBNs led to complex DNA damage and a specific increase of autophagic cell death. In addition, a significant radio-enhancement effect was obtained after the pre-conditioning of cells with a glutathione inhibitor before GBNs treatment and radiation exposure. Overall, our results provide additional information on the radio-enhancing properties of GBNs in the management of radioresistant HNSCC.

[Use of nanoparticles as radiosensitizing agents in radiotherapy: State of play]. / Utilisation de nanoparticules comme agent radiosensibilisant en radiothérapie : où en est-on ?

Nanomedicine has undergone significant development since the 2000s and it is only very recently that two metallic nanoparticles have emerged in clinical trials. The mechanism of these radiosensitizing agents is based on the presence of atoms with a high atomic number (Z) allowing a higher dose deposition into the tumor during irradiation. The first nanoparticle used in humans is NBTXR3, composed of hafnium (Z=79), with intratumor injection for the treatment of sarcoma. Another gadolinium-based nanoparticle (Z=64), AGuIX, has been used for intravenous injection in the treatment of brain metastases. The preliminary results are promising in terms of feasibility, safety and efficacy, as evidenced by the significant number of ongoing clinical trials. The upcoming challenges for the development of nanoparticles will be the targeting of cancer cells, their biodistribution into the body, their eventual toxicity and their industrial production. In the coming years, modalities of administration and optimal combinations with radiotherapy should be defined in connection with fundamental research.

[Long QT syndrome in children: analysis of the Lyon series]. / Syndrome du QT long congénital chez l'enfant. Analyse de la série lyonnaise.

Congenital long QT syndrome is a rare and serious disorder in children. In addition to the clinical and electrocardiographical diagnostic criteria, molecular biochemistry has identified six genes which are implicated in this pathology. Our study involved a retrospective analysis of 23 patients aged less than 21 with congenital long QT syndrome, followed up for an average of two years. Genotypes were obtained for all of the patients. There were unfortunately two deaths, one of which had a mutation in the SCN5A gene. The other patient had a double mutation of the SCN5A and KCNE2 genes. Symptomatic patients had QT and QTc intervals noticeably longer than the asymptomatic patients, although this difference was not shown to be significant. LQT3 patients as well as those with a double mutation were affected more severely because two of the three LQT3 patients and one of the two patients with a double mutation suffered a cardiac arrest. Three patients in our study showed no mutation. Nevertheless, two of them suffered a severe cardiac event. This confirms the limits of genetic diagnosis, which could be envisaged in all cases. All of the clinical and ECG data should be combined with the genetic analysis in order to confirm the diagnosis.

Abnormal cholesterol metabolism in imipramine-treated fibroblast cultures. Similarities with Niemann-Pick type C disease.

Addition of low-density lipoprotein (LDL) to cholesterol-deprived human skin fibroblast cultures treated by imipramine at a 20 microM concentration induced a significant intracellular accumulation of unesterified cholesterol. Intracytoplasmic inclusions were already visible by histochemical filipin staining after 2 h of LDL uptake and were progressively mobilized towards the perinuclear region within 24 h. At this concentration of the drug, the rate of proteolytic 125I-LDL hydrolysis was similar in treated and untreated cells. Treated cells maintained in lipoprotein-deficient medium showed no abnormality, indicating the exogenous origin of the accumulated sterol. Further, the drug induced a drastic dose-dependent impairment of LDL-stimulated cholesterol esterification, not related to an inhibition of acyl CoA:cholesterol acyltransferase, and a significant delay in down-regulation of de novo cholesterol synthesis. However, imipramine did not affect 25-hydroxycholesterol-mediated regulation of the two latter processes. These results resemble those observed in Niemann-Pick type C disease and suggest an impaired mobilization of LDL-derived cholesterol in imipramine-treated cells.

Free sphingoid bases in tissues from patients with type C Niemann-Pick disease and other lysosomal storage disorders.

The 20-fold increase of free sphingoid bases found in liver from a murine model of Niemann-Pick type C (NPC) combined to the NPC-like phenotype induced by addition of sphinganine to normal fibroblast cultures prompted us to investigate the potential involvement of these compounds in the human disease. The contents of sphingosine and sphinganine were measured in liver, spleen, brain and skin fibroblast cultures by a sensitive HPLC method. In liver and spleen from NPC patients, a 6- to 24-fold elevation of sphingosine and sphinganine already prominent at the fetal stage of the disease was observed, while no clear increase could be evidenced in brain tissue. A significant increase, not modulated by the intralysosomal content of free cholesterol, also occurred in skin fibroblast cultures. To investigate the specificity of these findings, other lysosomal storage disorders were studied. A striking accumulation was found in liver and spleen (24- to 36-fold) from patients with Niemann-Pick disease type A and B (sphingomyelinase-deficient forms), and in cerebral cortex of type A Niemann-Pick disease. A significant storage also occurred in Sandhoff disease, while several other sphingolipidoses showed a moderate elevation. In all cases but Sandhoff disease brain, the sphingosine/sphinganine ratio remained unchanged, suggesting that the accumulated free sphingoid bases derived from sphingolipid catabolism. Formation of complexes between sphingosine and the lipid material accumulated in lysosomes might be a general mechanism in lysosomal lipidoses. In NPC, however, an increase of free sphingoid bases disproportionate to the degree of lysosomal storage and a specific involvement of cultured fibroblasts suggested a more complex or combined mechanism.

Type C Niemann-Pick disease: spectrum of phenotypic variation in disruption of intracellular LDL-derived cholesterol processing.

To investigate biochemical heterogeneity within Niemann-Pick type C disease (NPC), the two most characteristic abnormalities, namely (1) kinetics of LDL-stimulated cholesteryl ester formation and (2) intravesicular accumulation of LDL-derived unesterified cholesterol, evaluated by histochemical filipin staining, were studied in cultured skin fibroblasts from a population of 125 NPC patients. Profound alterations (esterification rates less than 10% of normal, very numerous and intensely fluorescent cholesterol-filipin granules) were demonstrated in 86% of the cases, depicting the 'classical' NPC phenotype. The remaining cell lines showed a graded less severe impairment and more transient delay in the induction of LDL-mediated cholesteryl esterification, along with an attenuated accumulation of unesterified cholesterol. In particular, cells from a small group (7%) of patients, which have been individualized as representative of a 'variant' phenotype, showed only slight alterations of esterification, restricted to the early phase of LDL uptake and undistinguishable from those in heterozygotes. In these cells, an abnormal cytochemical distribution of LDL-derived cholesterol, although moderate, was still evident provided rigorous experimental conditions were followed. A third, less clearly individualized group (7%), differing from the classical phenotype mostly by higher rates of cholesteryl ester formation, has been designated as an 'intermediary' phenotype to reflect a more difficult diagnosis of such patients. These findings have an important bearing with regard to diagnosis and genetic counselling, although the significance of such a phenotypic variation in terms of genetic heterogeneity has still to be demonstrated. A given biochemical phenotype was however a constant observation within a family (14 pairs of siblings tested so far). The unique feature of LDL-cholesterol processing alterations in NPC has been further established from comparative studies in Wolman disease and I-cell disease, showing normal or different intracellular distribution of unesterified LDL-derived cholesterol in the latter disorders. Correlation between biochemical and clinical NPC phenotypes was only partial, but a correlation between the severity of alterations in cholesterol processing and sphingomyelin catabolism could be established.

Type C Niemann-Pick disease: a murine model of the lysosomal cholesterol lipidosis accumulates sphingosine and sphinganine in liver.

We have determined the levels of free sphingoid bases in livers of normal and cholesterol lipidotic Niemann-Pick type C mice. Hepatic sphingosine and sphinganine levels in affected mice (593 pmol/mg protein) were elevated more than 20-fold when compared to levels in age-matched normal mice (26 pmol/mg protein). Upon fractionation of mutant liver homogenates by differential centrifugation, most of the sphingoid bases sedimented with beta-hexosaminidase in the 9000 x g pellet. Co-sedimentation of sphingoid bases with a lighter beta-hexosaminidase peak in Percoll gradients suggests that these bases accumulate in lipid laden lysosomes. A cytosolic sphinganine kinase is the first enzyme in the degradative pathway of sphingoid base metabolism. Activity of this enzyme was partially deficient in crude mutant liver cytosolic extracts due to the presence of an inhibitory substance. Following molecular sieving of mutant cytosolic extracts on Sepharose 4B, sphinganine kinase, with normal levels of activity, was resolved from a complex higher-molecular-weight inhibitor fraction. The Km values for either sphinganine or ATP-Mg substrates with partially purified sphinganine kinase from normal and mutant mouse liver extracts, were similar. These findings indicate that accumulation of free sphingoid bases is not due to a direct inherent deficiency in the catalytic activity of sphinganine kinase. The possible cause and effect relationship between the accumulation of these endogenous hydrophobic amines and the lesion in intracellular cholesterol trafficking in Niemann-Pick type C disease is discussed.

[Cardiac troponin I and C: analytical comparison and clinical-biological interpretation of three troponins assays]. / Troponines I et T cardiaques: comparaison analytique et étude clinicobiologique de trois trousses de troponine.

Many assays 1(st), 2(nd) even 3(rd) generation are at present available to determine the concentration of cardiac troponin I and T. With the redefinition of upper reference value in the acute coronary syndromes, the aim of this study was to evaluate the clinical and analytical performance of 2 troponins assays: Troponin Ic 2(nd) generation (AccuTnI) on Access 2 of Beckman Coulter and Troponin Tc 3(rd)generation (Troponin T STAT) on Elecsys 2010 of Roche Diagnostics. The analytical performance observed with these 2 assays are accurate (analytical and functional sensitivity, repetability and reproductibility). Comparing each method with Dade Behring assay (Flex Troponine-I Cardiaque, TROP) on Dimension RxL, the correlation observed with AccuTnI kit on Access 2 can be put into the equation: AccuTnI = 1.08 (TnIc TROP) - 0.34, r = 0.99. On the contrary, it's more difficult to compare cTnI and cTnT. The study of decisonnal values indicated by Beckman Coulter for cTnI (0.04 microg/L at the 99 degrees percentil, 0.06 microg/L for a CV < or =10%) show a better specificity (76%) and predictive positive value (89%) with a sensitivity at 100% at 0.1 microg/L, fixed and used in the laboratory for its better agreement between sensibility / specificity and its imprecision below 10 %. For the cTnT values published by Roche Diagnostics (0.01 microg/L), at the 99 degrees percentil and 0.03 microg/L for a CV < or = 10%, the specificity is lower, so the decisionnal value 0.1 microg/L seems to be more suitable. During this study, few false positive and negative cTnT values have been observed, in patients with complex pathologies; this eventuality must be taken in consideration if clinical findings are not in good accordance with laboratory results.

Cellular and molecular portrait of eleven human glioblastoma cell lines under photon and carbon ion irradiation.

This study aimed to examine the cellular and molecular long-term responses of glioblastomas to radiotherapy and hadrontherapy in order to better understand the biological effects of carbon beams in cancer treatment. Eleven human glioblastoma cell lines, displaying gradual radiosensitivity, were irradiated with photons or carbon ions. Independently of p53 or O(6)-methylguanine-DNA methyltransferase(1) status, all cell lines responded to irradiation by a G2/M phase arrest followed by the appearance of mitotic catastrophe, which was concluded by a ceramide-dependent-apoptotic cell death. Statistical analysis demonstrated that: (i) the SF2(2) and the D10(3) values for photon are correlated with that obtained in response to carbon ions; (ii) regardless of the p53, MGMT status, and radiosensitivity, the release of ceramide is associated with the induction of late apoptosis; and (iii) the appearance of polyploid cells after photon irradiation could predict the Relative Biological Efficiency(4) to carbon ions. This large collection of data should increase our knowledge in glioblastoma radiobiology in order to better understand, and to later individualize, appropriate radiotherapy treatment for patients who are good candidates.

Preparation of an anti-acid sphingomyelinase monoclonal antibody for the quantitative determination and polypeptide analysis of lysosomal sphingomyelinase in fibroblasts from normal and Niemann-Pick type A patients.

An anti-acid sphingomyelinase monoclonal antibody has been prepared using an in vitro booster technique. The antigen, acid sphingomyelinase, was purified from human placentas by sequential chromatographic steps in the presence of the non-ionic detergent Nonidet P40. This monoclonal antibody (MAB 236) precipitates specifically the enzyme activity by immunoadsorption techniques and presents the same specificity to normal and mutated sphingomyelinase in Niemann-Pick type A patients. MAB 236 is the first antibody able to precipitate the protein in the presence of detergent thereby permitting the quantitative determination of normal and mutated sphingomyelinase in tissue and cell extracts. Polypeptide analysis and quantitative determination experiments using this monoclonal antibody showed no difference between patients and normal controls.

Ceramide induces activation of the mitochondrial/caspases pathway in Jurkat and SCC61 cells sensitive to gamma-radiation but activation of this sequence is defective in radioresistant SQ20B cells.

PURPOSE: To clarify the molecular mechanisms leading to radiation-induced apoptosis or resistance, the kinetics (1-48 h) and sequence of events triggered in response to 10 Gy irradiation were investigated in three cell lines displaying a gradient of sensitivity to 7-rays. MATERIALS AND METHODS: Ceramide levels were measured by high performance liquid chromatography (HPLC). Mitochondrial function was evaluated in terms of transmembrane potential (delta(psi)m), reactive oxygen species (ROS) and glutathione levels analysed by flow cytometry or HPLC. Caspase activation was assessed by immunoblotting, and apoptosis by flow cytometry. RESULTS: In Jurkat radiosensitive cells and SCC61 adherent cells with intermediate radiosensitivity, the degree of delayed ceramide release was directly related to their propensity to undergo apoptosis. Transduction of the death signal was mediated by a drop in delta(psi)m and glutathione levels, ROS accumulation and activation of effector caspases. Experiments conducted with caspase inhibitors, bongkrekic acid, or DL-PDMP indicated that ceramide triggers mitochondrial collapse, followed by the activation of caspases-9, -8 and -3, and poly(ADP-ribose)polymerase cleavage. In SQ20B radioresistant cells, gamma-radiation did not induce ceramide generation or subsequent activation of the mitochondrial/caspase apoptotic pathway. CONCLUSIONS: Ceramide appears to be a determining factor in the commitment phase of radiation-induced apoptosis. When ceramide is not generated, the whole pathway is ineffective and resistance to apoptosis may result.

Comparison of methods used to study cell death in an adherent tumoral cell line with moderate clonogenic radiosensitivity.

Our objective was to compare different methods for studying programmed cell death in adherent H460 non-small lung cancer cells of moderate clonogenic radiosensitivity. The major effect of gamma-radiation was found to be the release of cells from the substratum. The different methods gave complementary and unexpected information: a) with the TUNEL method, a few non-apoptotic cells were found in the culture medium; b) with the flow cytometry after propidium iodide labeling, some hypodiploid cells which remained attached to the substratum were apoptotic, as demonstrated by the effect of a caspase inhibitor; c) with the annexin V labeling, the detached cells were demonstrated either necrotic or very late apoptotic; d) the mitochondria transmembrane potential (deltapsim), measurements demonstrated that the mitochondria were implicated in cell death induced by gamma-radiation. These data illustrate the need to use several complementary methods in the study of apoptosis in adherent cells exposed to gamma-radiation.

[HDL-cholesterol: role of its dosage in the assessment of cardiovascular risk]. / HDL-cholestérol: place de son dosage dans l'évaluation d'un risque cardiovasculaire.

The interest of HDL-cholesterol (HDLC) to evaluate a cardiovascular risk has been studied since many years. According to Framingham Heart studies, this factor is inversely correlated to a future ischaemic heart disease. At high level, HDLC is considered as a cardiovascular protecting factor, and is known since few years as "good cholesterol". In the year 2000, the ANAES (Agence nationale de l'accréditation et évaluation en santé) has redefined the role of HDLC in the exploration of dyslipidaemia. In the case of a cardiovascular-risk history, HDLC, with total cholesterol, triglycerides, and LDL-cholesterol (by Friedewald method) will be analyzed. Usually, HDLC is not very accessible to conventional treatments. So, according to ANAES, the treatment of dyslipidaemia will be based on LDL-cholesterol levels only. Nevertheless, HDLC is a major lipid factor to evaluate a cardiovascular risk. The object of this review is, on one hand, to situate HDLC in the evaluation of cardiovascular risk, by showing its key role in lipid metabolism, and, on the other hand, to report the main direct assays of this parameter.

The use of theranostic gadolinium-based nanoprobes to improve radiotherapy efficacy.

A new efficient type of gadolinium-based theranostic agent (AGuIX®) has recently been developed for MRI-guided radiotherapy (RT). These new particles consist of a polysiloxane network surrounded by a number of gadolinium chelates, usually 10. Owing to their small size (<5 nm), AGuIX typically exhibit biodistributions that are almost ideal for diagnostic and therapeutic purposes. For example, although a significant proportion of these particles accumulate in tumours, the remainder is rapidly eliminated by the renal route. In addition, in the absence of irradiation, the nanoparticles are well tolerated even at very high dose (10 times more than the dose used for mouse treatment). AGuIX particles have been proven to act as efficient radiosensitizers in a large variety of experimental in vitro scenarios, including different radioresistant cell lines, irradiation energies and radiation sources (sensitizing enhancement ratio ranging from 1.1 to 2.5). Pre-clinical studies have also demonstrated the impact of these particles on different heterotopic and orthotopic tumours, with both intratumoural or intravenous injection routes. A significant therapeutical effect has been observed in all contexts. Furthermore, MRI monitoring was proven to efficiently aid in determining a RT protocol and assessing tumour evolution following treatment. The usual theoretical models, based on energy attenuation and macroscopic dose enhancement, cannot account for all the results that have been obtained. Only theoretical models, which take into account the Auger electron cascades that occur between the different atoms constituting the particle and the related high radical concentrations in the vicinity of the particle, provide an explanation for the complex cell damage and death observed.

[From the carbon track to therapeutic efficiency of hadrontherapy]. / De la trace des ions carbone à l'efficacité thérapeutique de l'hadronthérapie.

Carbon ions, thanks to their relative biological effectiveness much higher than that of photons and protons and their ballistic characteristics similar to those of protons, can effectively treat radioresistant tumours. The reasons for this increased efficiency are found in the microdosimetric and radiobiological features of ions. The energy deposit or linear energy transfer increases along the range and reaches a very high level at the end producing the Bragg peak, where the linear energy transfer is about hundred times higher than that of photons. These massive energy deposits create multiple DNA lesions that are difficult to repair. DNA repair is associated with longer blockage of the cell cycle and more frequent chromosomal aberrations that are lethal to cells. The types of cell death are identical to those triggered in response to photon irradiation, but the response is earlier and more important at equivalent physical dose. Radiobiological differences between carbon ions and photons have been studied for some years and many aspects remain to be explored. In general, these phenomena tend to reduce the differences of radiosensitivity among different tissues. It is therefore in situation where tumours are relatively radioresistant compared to healthy tissue, that carbon ions must be used and not in the opposite situations where the fractionation of low linear energy transfer radiation is sufficient to provide the necessary differential effect to cure the tumour.

Inhibition of heat shock protein 27 (HspB1) tumorigenic functions by peptide aptamers.

Human heat shock protein 27 (Hsp27, HspB1) is an anti-apoptotic protein characterized for its tumorigenic and metastatic properties, and now referenced as a major therapeutic target in many types of cancer. Hsp27 biochemical properties rely on a structural oligomeric and dynamic organization. Downregulation by small interfering RNA or inhibition with dominant-negative mutant have proven their efficiency to counteract the anti-apoptotic and protective properties of Hsp27. In this study, we report the isolation and characterization of Hsp27-targeted molecules interfering with its structural organization. Using the peptide aptamer (PA) strategy, we isolated PAs that specifically interact with Hsp27 and not with the other members of the small heat shock protein family. In mammalian cell cultures, PAs expression perturbed the dimerization and oligomerization of Hsp27, and acted as negative regulators of the anti-apoptotic and cytoprotective activities of this protein. Further studies analyzing SQ20B cell xenografts in immunocompromised mice showed that PAs strongly reduced tumor development through cell cycle arrest. Our data suggest that PAs could provide a potential tool to develop strategies for the discovery of Hsp27 chemical inhibitors.

Spectrum of pathogenic mutations and associated polymorphisms in a cohort of 44 unrelated patients with long QT syndrome.

Long QT syndrome (LQTS) is a rare and clinically heterogeneous inherited disorder characterized by a long QT interval on the electrocardiogram, increased risk of syncope and sudden death caused by arrhythmias. This syndrome is mostly caused by mutations in genes encoding various cardiac ion channels. The clinical heterogeneity is usually attributed to variable penetrance. One of the reasons for this variability in expression could be the coexistence of common single nucleotide polymorphisms (SNPs) on LQTS-causing genes and/or unknown genes. Some synonymous and nonsynonymous exonic SNPs identified in LQTS-causing genes may have an effect on the cardiac repolarization process and modulate the clinical expression of a latent LQTS pathogenic mutation. We report the molecular pattern of 44 unrelated patients with LQTS using denaturing high-performance liquid chromatography analysis of the KCNQ1, KCNH2, SCN5A, KCNE1 and KCNE2 genes. Forty-five disease-causing mutations (including 24 novel ones) were identified in this cohort. Most of our patients (84%) showed complex molecular pattern with one mutation (and even two for four patients) associated with several SNPs located in several LQTS genes.