Sofosbuvir and Velpatasvir for HCV Genotype 2 and 3 Infection

BACKGROUND: In phase 2 trials, treatment with the combination of the nucleotide polymerase inhibitor sofosbuvir and the NS5A inhibitor velpatasvir resulted in high rates of sustained virologic response in patients chronically infected with hepatitis C virus (HCV) genotype 2 or 3. METHODS: We conducted two randomized, phase 3, open-label studies involving patients who had received previous treatment for HCV genotype 2 or 3 and those who had not received such treatment, including patients with compensated cirrhosis. In one trial, patients with HCV genotype 2 were randomly assigned in a 1:1 ratio to receive sofosbuvir-velpatasvir, in a once-daily, fixed-dose combination tablet (134 patients), or sofosbuvir plus weight-based ribavirin (132 patients) for 12 weeks. In a second trial, patients with HCV genotype 3 were randomly assigned in a 1:1 ratio to receive sofosbuvir-velpatasvir for 12 weeks (277 patients) or sofosbuvir-ribavirin for 24 weeks (275 patients). The primary end point for the two trials was a sustained virologic response at 12 weeks after the end of therapy. RESULTS: Among patients with HCV genotype 2, the rate of sustained virologic response in the sofosbuvir-velpatasvir group was 99% (95% confidence interval [CI], 96 to 100), which was superior to the rate of 94% (95% CI, 88 to 97) in the sofosbuvir-ribavirin group (P=0.02). Among patients with HCV genotype 3, the rate of sustained virologic response in the sofosbuvir-velpatasvir group was 95% (95% CI, 92 to 98), which was superior to the rate of 80% (95% CI, 75 to 85) in the sofosbuvir-ribavirin group (P CONCLUSIONS: Among patients with HCV genotype 2 or 3 with or without previous treatment, including those with compensated cirrhosis, 12 weeks of treatment with sofosbuvir-velpatasvir resulted in rates of sustained virologic response that were superior to those with standard treatment with sofosbuvir-ribavirin. (Funded by Gilead Sciences; ASTRAL-2 ClinicalTrials.gov number, NCT02220998; and ASTRAL-3, NCT02201953.)

Identification of germline alterations of the mad homology 2 domain of SMAD3 and SMAD4 from the Ontario site of the breast cancer family registry (CFR)

Abstract Introduction A common feature of neoplastic cells is that mutations in SMADs can contribute to the loss of sensitivity to the anti-tumor effects of transforming growth factor-β (TGF-β). However, germline mutation analysis of SMAD3 and SMAD4, the principle substrates of the TGF-β signaling pathway, has not yet been conducted in breast cancer. Thus, it is currently unknown whether germline SMAD3 and SMAD4 mutations are involved in breast cancer predisposition. Methods We performed mutation analysis of the highly conserved mad-homology 2 (MH2) domains for both genes in genomic DNA from 408 non-BRCA1/BRCA2 breast cancer cases and 710 population controls recruited by the Ontario site of the breast cancer family registry (CFR) using denaturing high-performance liquid chromatography (DHPLC) and direct DNA sequencing. The results were interpreted in several ways. First, we adapted nucleotide diversity analysis to quantitatively assess whether the frequency of alterations differ between the two genes. Next, in silico tools were used to predict variants' effect on domain function and mRNA splicing. Finally, 37 cases or controls harboring alterations were tested for aberrant splicing using reverse-transcription polymerase chain reaction (PCR) and real-time PCR statistical comparison of germline expressions by non-parametric Mann-Whitney test of independent samples. Results We identified 27 variants including 2 novel SMAD4 coding variants c.1350G > A (p.Gln450Gln), and c.1701A > G (p.Ile525Val). There were no inactivating mutations even though c.1350G > A was predicted to affect exonic splicing enhancers. However, several additional findings were of note: 1) nucleotide diversity estimate for SMAD3 but not SMAD4 indicated that coding variants of the MH2 domain were more infrequent than expected; 2) in breast cancer cases SMAD3 was significantly over-expressed relative to controls (P A was associated with elevated germline expression (> 5-fold); 3) separate analysis using tissue expression data showed statistically significant over-expression of SMAD3 and SMAD4 in breast carcinomas. Conclusions This study shows that inactivating germline alterations in SMAD3 and SMAD4 are rare, suggesting a limited role in driving tumorigenesis. Nevertheless, aberrant germline expressions of SMAD3 and SMAD4 may be more common in breast cancer than previously suspected and offer novel insight into their roles in predisposition and/or progression of breast cancer

Failure of human rhombic lip differentiation underlies medulloblastoma formation

Medulloblastoma (MB) comprises a group of heterogeneous paediatric embryonal neoplasms of the hindbrain with strong links to early development of the hindbrain 1–4. Mutations that activate Sonic hedgehog signalling lead to Sonic hedgehog MB in the upper rhombic lip (RL) granule cell lineage 5–8. By contrast, mutations that activate WNT signalling lead to WNT MB in the lower RL 9,10. However, little is known about the more commonly occurring group 4 (G4) MB, which is thought to arise in the unipolar brush cell lineage 3,4. Here we demonstrate that somatic mutations that cause G4 MB converge on the core binding factor alpha (CBFA) complex and mutually exclusive alterations that affect CBFA2T2, CBFA2T3, PRDM6, UTX and OTX2. CBFA2T2 is expressed early in the progenitor cells of the cerebellar RL subventricular zone in Homo sapiens, and G4 MB transcriptionally resembles these progenitors but are stalled in developmental time. Knockdown of OTX2 in model systems relieves this differentiation blockade, which allows MB cells to spontaneously proceed along normal developmental differentiation trajectories. The specific nature of the split human RL, which is destined to generate most of the neurons in the human brain, and its high level of susceptible EOMES +KI67 + unipolar brush cell progenitor cells probably predisposes our species to the development of G4 MB

Missense Variants of Uncertain Significance (VUS) Altering the Phosphorylation Patterns of BRCA1 and BRCA2

Mutations in BRCA1 and BRCA2 are responsible for a large proportion of breast-ovarian cancer families. Protein-truncating mutations have been effectively used in the clinical management of familial breast cancer due to their deleterious impact on protein function. However, the majority of missense variants identified throughout the genes continue to pose an obstacle for predictive informative testing due to low frequency and lack of information on how they affect BRCA1/2 function. Phosphorylation of BRCA1 and BRCA2 play an important role in their function as regulators of DNA repair, transcription and cell cycle in response to DNA damage but whether missense variants of uncertain significance (VUS) are able to disrupt this important process is not known. Here we employed a novel approach using NetworKIN which predicts in vivo kinasesubstrate relationship, and evolutionary conservation algorithms SIFT, PolyPhen and Align-GVGD. We evaluated whether 191 BRCA1 and 43 BRCA2 VUS from the Breast Cancer Information Core (BIC) database can functionally alter the consensus phosphorylation motifs and abolish kinase recognition and binding to sites known to be phosphorylated in vivo. Our results show that 13.09% (25/191) BRCA1 and 13.95% (6/43) BRCA2 VUS altered the phosphorylation of BRCA1 and BRCA2. We highlight six BRCA1 (K309T, S632N, S1143F, Q1144H, Q1281P, S1542C) and three BRCA2 (S196I, T207A, P3292L) VUS as potentially clinically significant. These occurred rarely (n,2 in BIC), mutated evolutionarily conserved residues and abolished kinase binding to motifs established in the literature involved in DNA repair, cell cycle regulation, transcription or response to DNA damage. Additionally in vivo phosphorylation sites identified via through-put methods are also affected by VUS and are attractive targets for studying their biological and functional significance. We propose that rare VUS affecting phosphorylation may be a novel and important mechanism for which BRCA1 and BRCA2 functions are disrupted in breast cancer

Identification of Germline Alterations in the Mad-homology 2 (MH2) Domain of SMAD3 and SMAD4 In Breast Cancer Susceptibility

A feature of neoplastic cells is that mutations in the key intermediates of TGF-β signaling contribute to the loss of sensitivity to its anti-tumor effects. The role of SMAD3 and SMAD4 germline mutations in breast cancer predisposition is currently unclear. To address this, mutation analysis of the Mad-Homology 2 domains in 408 breast cancer cases and 710 controls recruited by the Breast Cancer Family Registry (BCFR) was performed using Denaturing High-Pressure Liquid Chromatography. This study identified 23 distinct intronic variants, and three coding variants c.1214T>C, c.1478G>A, and c.1701A>G in SMAD4. No aberrant splicing was observed, but qPCR analysis and tissue expression data showed significantly elevated SMAD3 expression relative to controls (pA from a familial breast cancer case showed a 5-fold expression change. Taken together, inactivating alterations are not driving tumorigenesis. Rather, aberrant germline expression provides novel insight into SMAD3 and SMAD4’s roles in breast cancer predisposition.MAS

Structural and mechanical properties of AlCrN thin films

Ce travail est consacré à l'étude systématique de films Al1-xCrxNy depuis leur production jusqu'à leur caractérisation structural et mécanique. Les films ont été synthétises à l'aide de deux systèmes PVD : pulvérisation par faisceau d'ion (IBS) et pulvérisation magnétron (MS). La composition des films est étudiée par EDXS et RBS, cette derrière permettant l'analyse des éléments légers tel que l azote. La majorité des compositions sont sous- stoechiométriques avec une concentration en azote (y < 1) décroissant continûment lorsque la concentration de Chrome (x) croît. Ce résultat a été confirmé par des simulations TRIM. La structure et la morphologie des films (épaisseur, phase, texture, taille des nano-grains) ont été étudiées essentiellement par diffraction et réflectométrie des rayons X. Les contraintes résiduelles (global, in-situ et intra granulaire) ont été étudiées par la méthode de la courbure et diffraction X. Les résultats montrent l'importance tant de la composition (x, y) des films que du type de substrat. Deux techniques sensibles (l'acoustique picoseconde et la diffusion brillouin) ont été utilisées pour l'étude des propriétés élastiques et ont révélé, une transition abrupte à x = 0.4. La technique de nano-indentation a été appliquée pour analyser le module d'indentation et la dureté. Des essais de tribologie ont permis de compléter l'étude par la détermination des coefficients de frottement (COF).This work is dedicated to the systematical study of Al1-xCrxNy coatings starting from coating processes to mechanical properties through composition and phase structure investigations. Al1-xCrxNy thin films have been synthesized by two PVD systems: ion-beam sputtering (IBS) and magnetron sputtering (MS). Composition of the coatings is studied by both EDXS and RBS, this latter being very sensitive to light elements such as nitrogen. Most compositions are sub - stoichiometric with Nitrogen content (y < 1) decreasing continuously when Chromium content (x) increases. This result has been confirmed by the TRIM s simulation. Structure and morphology of the films which include thickness, phases and texture, Scherrer s nano grain - size are analyzed mainly by x - ray techniques such as diffraction and reflectometry. Residual stresses including global residual stresses, real - time in - situ global residual stresses, thermal stresses and in - grain residual stresses are studied using substrate curvature and XRD methods. It emphasizes the importance of considering both composition (x, y) and substrate type in the study of mechanical properties. Two sensitive methods (Picoseconds ultrasonic and Brillouin Light Scattering) were used to study elasticity of the AlCrN films and revealed a sharp transition at x = 0.4. Nanoindentation technique was applied for analyze indentation modulus and hardness of our Al1-xCrxNy films and furthermore tribological testing for investigating coefficients of friction (COF) of the films deposited on stainless steels by pin on - disk method.POITIERS-BU Sciences (861942102) / SudocSudocFranceF

Multiple sequence alignment demonstrating phylogenetic conservation of the three biologically characterized phosphorylated BRCA2 residues affected by missense variants of unknown clinical significance.

<p>Multiple sequence alignment demonstrating phylogenetic conservation of the three biologically characterized phosphorylated BRCA2 residues affected by missense variants of unknown clinical significance.</p

Figure 1

<p><b>a.</b> Summary of phosphorylation sites studied in BRCA1. Residues in green represent <i>in vivo</i> phosphorylation sites have been biologically characterized in the literature. Residues in red represent <i>in vivo</i> phosphorylation sites identified via throughput methods where biological functions have not yet been determined. <b>b.</b> Summary of phosphorylation sites studied in BRCA2. Residues in green represent <i>in vivo</i> phosphorylation sites that have been biologically characterized in the literature. Residues in red represent <i>in vivo</i> phosphorylation sites identified via throughput methods where biological functions have not yet been determined.</p

NetworKIN analysis of VUS affecting biologically uncharacterized phosphorylation sites in BRCA1 and BRCA2.

<p>In <b>bold</b> are BRCA1 mutations that fall within an experimentally identified but biologically uncharacterized phosphorylation site. ^aThe position and change at the amino acids specified by the missense variant is as reported in the BIC database. ^bThe nucleotide change conforms to the HGVS nomenclature. ^cSNP IDs correspond to the dbSNP database <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0062468#pone.0062468-Sherry1" target="_blank">[73]</a> SNP identifiers. ^dFrequency represents the number of times reported in the BIC database. ^eThe ten-residue long biologically uncharacterized kinase recognition motifs are shown. The biologically uncharacterized Serine (S), and threonine (T) residues shown to be phosphorylated by NetworKIN are underlined. * Sites that retained a score but was considered to be “abolished” due to score falling below 5 with the presence of the VUS.</p