Regulatory Provision of the Passenger's Rights in the System of Warranties of their Compliance and Realization

The article deals with topical issues of securing the rights of the passenger. As violations of this kind of rights are frequent and the means of protection of their rights, except for air transport, are virtually absent under the transport law of Ukraine, except for the possibility of compensation for non-pecuniary damage, the presence and amount of which is determined by court procedure, there is an urgent need to determine and regulate this civil rights, how it is under EU law on new civilizational approaches; securing regulatory safeguards to enforce and exercise their rights. The authors conclude that according to the legislation of Ukraine there are practically no guarantees for the exercise of the rights of the passenger, which is unacceptable at this stage of civilization development. As the mechanism for securing this kind of civil rights under EU law, which is based on the concept of creating a competitive environment, regulating and securing the rights of passengers, establishing negative consequences for the carrier in the event of a violation of the passenger's rights in the form of compensation amounts, despite the fact that he is debatable and imperfect, however, it can be implemented into the national legislation of Ukraine. In addition to the right to require the carrier to deliver it to its destination, the basic rights of the passenger must be legally enshrined and guaranteed: security, human rights, mobility, non-discrimination, assistance in transportation, accessibility, full and reliable information, on the use of remedies, on compensation in case of violation of rights, etc

Homologous Recombination Deficiency Across Subtypes of Primary Breast Cancer

Homologous recombination; Primary breast cancerRecombinació homòloga; Càncer de mama primariRecombinación homóloga; Cáncer de mama primarioPURPOSE Homologous recombination deficiency (HRD) is highly prevalent in triple-negative breast cancer (TNBC) and associated with response to PARP inhibition (PARPi). Here, we studied the prevalence of HRD in non-TNBC to assess the potential for PARPi in a wider group of patients with breast cancer. METHODS HRD status was established using targeted gene panel sequencing (360 genes) and BRCA1 methylation analysis of pretreatment biopsies from 201 patients with primary breast cancer in the phase II PETREMAC trial (ClinicalTrials.gov identifier: NCT02624973). HRD was defined as mutations in BRCA1, BRCA2, BRIP1, BARD1, or PALB2 and/or promoter methylation of BRCA1 (strict definition; HRD-S). In secondary analyses, a wider definition (HRD-W) was used, examining mutations in 20 additional genes. Furthermore, tumor BRCAness (multiplex ligation-dependent probe amplification), PAM50 subtyping, RAD51 nuclear foci to test functional HRD, tumor-infiltrating lymphocyte (TIL), and PD-L1 analyses were performed. RESULTS HRD-S was present in 5% of non-TNBC cases (n = 9 of 169), contrasting 47% of the TNBC tumors (n = 15 of 32). HRD-W was observed in 23% of non-TNBC (n = 39 of 169) and 59% of TNBC cases (n = 19 of 32). Of 58 non-TNBC and 30 TNBC biopsies examined for RAD51 foci, 4 of 4 (100%) non-TNBC and 13 of 14 (93%) TNBC cases classified as HRD-S had RAD51 low scores. In contrast, 4 of 17 (24%) non-TNBC and 15 of 19 (79%) TNBC biopsies classified as HRD-W exhibited RAD51 low scores. Of nine non-TNBC tumors with HRD-S status, only one had a basal-like PAM50 signature. There was a high concordance between HRD-S and either BRCAness, high TIL density, or high PD-L1 expression (each P < .001). CONCLUSION The prevalence of HRD in non-TNBC suggests that therapy targeting HRD should be evaluated in a wider breast cancer patient population. Strict HRD criteria should be implemented to increase diagnostic precision with respect to functional HRD.Supported by unrestricted grants from The K.G. Jebsen Foundation [SKGJ-MED-020 to H.P.E., S.K., P.E.L.], Helse Vest [912008 to P.E.L.], The Norwegian Research Council [273354 to P.E.L.] and The Norwegian Cancer Society [190281-2017 to S.K., 190275-2017 to P.E.L.]. Research funding was provided by Grieg Foundation (to H.P.E. and T.A.), Helse Vest [912252; Clinical researcher fellowship to H.P.E.], Generalitat de Catalunya [PERIS, SLT017/20/000081 to A.H.R.], “la Caixa” Foundation and European Institute of Innovation and Technology/Horizon 2020 [LCF/TR/CC19/52470003 to A.L.G.], Asociación Española Contra el Cáncer [AECC, INVES20095LLOP to A.L.G.], Generalitat de Catalunya (PERIS Fellowship SLT002/16/00477 to A.L.G.) ERA PerMed [2019-215 to V.S.] and Miguel Servet fellowship (CPII19/00033 to V.S.). Additional funding and study medication was provided by Illumina [grant number 9529854], Pfizer [WI206347] and AstraZeneca [ESR-14-10077] to H.P.E., S.K., P.E.L

GSK3 alpha and GSK3 beta phosphorylate arc and regulate its degradation

The selective and neuronal activity-dependent degradation of synaptic proteins appears to be crucial for long-term synaptic plasticity. One such protein is activity-regulated cytoskeleton-associated protein (Arc), which regulates the synaptic content of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPAR), excitatory synapse strength and dendritic spine morphology. The levels of Arc protein are tightly regulated, and its removal occurs via proteasome-mediated degradation that requires prior ubiquitination. Glycogen synthase kinases α and β (GSK3α, GSKβ; collectively named GSK3α/β) are serine-threonine kinases with abundant expression in the central nervous system. Both GSK3 isozymes are tonically active under basal conditions, but their activity is regulated by intra- and extracellular factors, intimately involved in neuronal activity. Similar to Arc, GSK3α and GSK3β contribute to synaptic plasticity and the structural plasticity of dendritic spines. The present study identified Arc as a GSK3α/β substrate and showed that GSKβ promotes Arc degradation under conditions that induce de novo Arc synthesis. We also found that GSK3α/β inhibition potentiated spine head thinning that was caused by the prolonged stimulation of N-methyl-D-aspartate receptors (NMDAR). Furthermore, overexpression of Arc mutants that were resistant to GSK3β-mediated phosphorylation or ubiquitination resulted in a stronger reduction of dendritic spine width than wildtype Arc overexpression. Thus, GSK3β terminates Arc expression and limits its effect on dendritic spine morphology. Taken together, the results identify GSK3α/β-catalyzed Arc phosphorylation and degradation as a novel mechanism for controlling the duration of Arc expression and function

Alternative splicing affecting the sh3a domain controls the binding properties of intersectin 1 in neurons

a b s t r a c t Intersectin 1 (ITSN1) is a conserved adaptor protein implicated in endocytosis, regulation of actin cytoskeleton rearrangements and mitogenic signaling. Its expression is characterized by multiple alternative splicing. Here we show neuron-specific expression of ITSN1 isoforms containing exon 20, which encodes five amino acid residues in the first SH3 domain (SH3A). In vitro binding experiments demonstrated that inclusion of exon 20 changes the binding properties of the SH3A domain. Endocytic proteins dynamin 1 and synaptojanin 1 as well as GTPase-activating protein CdGAP bound the neuron-specific variant of the SH3A domain with higher affinity than ubiquitously expressed SH3A. In contrast, SOS1, a guanine nucleotide exchange factor for Ras, and the ubiquitin ligase Cbl mainly interact with the ubiquitously expressed isoform. These results demonstrate that alternative splicing leads to the formation of two pools of ITSN1 with potentially different properties in neurons, affecting ITSN1 function as adaptor protein. Ó 2008 Elsevier Inc. All rights reserved. Intersectin 1 (ITSN1) is an evolutionarily conserved adaptor protein involved in clathrin-mediated endocytosis, apoptosis, signal transduction, and regulation of cytoskeletal rearrangements ITSN1 has been shown to interact with several essential endocytic proteins, including dynamin, epsin, Eps15, and synaptojanin Previously, we found seven additional splice variants of the human and mouse ITSN1 genes In view of the potential link between alternative exon 20 usage and the binding properties of ITSN1, we have investigated the interaction of two SH3A domain splice variants with the main ITSN1 protein partners. The results presented here highlight differences in the binding properties of alternatively spliced SH3A domains of ITSN1 and showed how alternative splicing can regulate the functions of ITSN1 as adaptor protein. Materials and methods Expression constructs. The cDNA sequences corresponding to the proline-rich region (PRD) of dynamin 1 (residues 742-851, GenBank Accession No. NP_0010053360), SOS1-IsfI (residues 0006-291X/$ -see front matter

Homologous Recombination Deficiency Across Subtypes of Primary Breast Cancer

Purpose - Homologous recombination deficiency (HRD) is highly prevalent in triple-negative breast cancer (TNBC) and associated with response to PARP inhibition (PARPi). Here, we studied the prevalence of HRD in non-TNBC to assess the potential for PARPi in a wider group of patients with breast cancer. Methods - HRD status was established using targeted gene panel sequencing (360 genes) and BRCA1 methylation analysis of pretreatment biopsies from 201 patients with primary breast cancer in the phase II PETREMAC trial (ClinicalTrials.gov identifier: NCT02624973). HRD was defined as mutations in BRCA1, BRCA2, BRIP1, BARD1, or PALB2 and/or promoter methylation of BRCA1 (strict definition; HRD-S). In secondary analyses, a wider definition (HRD-W) was used, examining mutations in 20 additional genes. Furthermore, tumor BRCAness (multiplex ligation-dependent probe amplification), PAM50 subtyping, RAD51 nuclear foci to test functional HRD, tumor-infiltrating lymphocyte (TIL), and PD-L1 analyses were performed. Results - HRD-S was present in 5% of non-TNBC cases (n = 9 of 169), contrasting 47% of the TNBC tumors (n = 15 of 32). HRD-W was observed in 23% of non-TNBC (n = 39 of 169) and 59% of TNBC cases (n = 19 of 32). Of 58 non-TNBC and 30 TNBC biopsies examined for RAD51 foci, 4 of 4 (100%) non-TNBC and 13 of 14 (93%) TNBC cases classified as HRD-S had RAD51 low scores. In contrast, 4 of 17 (24%) non-TNBC and 15 of 19 (79%) TNBC biopsies classified as HRD-W exhibited RAD51 low scores. Of nine non-TNBC tumors with HRD-S status, only one had a basal-like PAM50 signature. There was a high concordance between HRD-S and either BRCAness, high TIL density, or high PD-L1 expression (each P Conclusion - The prevalence of HRD in non-TNBC suggests that therapy targeting HRD should be evaluated in a wider breast cancer patient population. Strict HRD criteria should be implemented to increase diagnostic precision with respect to functional HRD

ramr: an R/Bioconductor package for detection of rare aberrantly methylated regions

Motivation: With recent advances in the field of epigenetics, the focus is widening from large and frequent disease- or phenotype-related methylation signatures to rare alterations transmitted mitotically or transgenerationally (constitutional epimutations). Merging evidence indicate that such constitutional alterations, albeit occurring at a low mosaic level, may confer risk of disease later in life. Given their inherently low incidence rate and mosaic nature, there is a need for bioinformatic tools specifically designed to analyze such events. Results: We have developed a method (ramr) to identify aberrantly methylated DNA regions (AMRs). ramr can be applied to methylation data obtained by array or next-generation sequencing techniques to discover AMRs being associated with elevated risk of cancer as well as other diseases. We assessed accuracy and performance metrics of ramr and confirmed its applicability for analysis of large public datasets. Using ramr we identified aberrantly methylated regions that are known or may potentially be associated with development of colorectal cancer and provided functional annotation of AMRs that arise at early developmental stages.publishedVersio

Implication of the APP gene in intellectual abilities

Background: Cognitive functions are highly heritable and polygenic, though the source of this genetic influence is unclear. On the neurobiological level, these functions rely on effective neuroplasticity, in which the activity-regulated cytoskeleton associated protein (ARC) plays an essential role. Objectives: To examine whether the ARC gene complex may contribute to the genetic components of intellectual function given the crucial role of ARC in brain plasticity and memory formation. Methods: The ARC complex was tested for association with intelligence (IQ) in children from the Avon Longitudinal Study of Parents and Children (ALSPAC, N = 5,165). As Alzheimer’s disease (AD) shares genetics with cognitive functioning, the association was followed up in an AD sample (17,008 cases, 37,154 controls). Results: The ARC complex revealed association with verbal and total IQ (empirical p = 0.027 and 0.041, respectively) in the ALSPAC. The strongest single variant signal (rs2830077; empirical p = 0.018), within the APP gene, was confirmed in the AD sample (p = 2.76E-03). Functional analyses of this variant showed its preferential binding to the transcription factor CP2. Discussion: This study implicates APP in childhood IQ. While follow-up studies are needed, this observation could help elucidate the etiology of disorders associated with cognitive dysfunction, such as AD

GSK3α and GSK3β Phosphorylate Arc and Regulate its Degradation

The selective and neuronal activity-dependent degradation of synaptic proteins appears to be crucial for long-term synaptic plasticity. One such protein is activity-regulated cytoskeleton-associated protein (Arc), which regulates the synaptic content of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPAR), excitatory synapse strength and dendritic spine morphology. The levels of Arc protein are tightly regulated, and its removal occurs via proteasome-mediated degradation that requires prior ubiquitination. Glycogen synthase kinases α and β (GSK3α, GSKβ; collectively named GSK3α/β) are serine-threonine kinases with abundant expression in the central nervous system. Both GSK3 isozymes are tonically active under basal conditions, but their activity is regulated by intra- and extracellular factors, intimately involved in neuronal activity. Similar to Arc, GSK3α and GSK3β contribute to synaptic plasticity and the structural plasticity of dendritic spines. The present study identified Arc as a GSK3α/β substrate and showed that GSKβ promotes Arc degradation under conditions that induce de novo Arc synthesis. We also found that GSK3α/β inhibition potentiated spine head thinning that was caused by the prolonged stimulation of N-methyl-D-aspartate receptors (NMDAR). Furthermore, overexpression of Arc mutants that were resistant to GSK3β-mediated phosphorylation or ubiquitination resulted in a stronger reduction of dendritic spine width than wildtype Arc overexpression. Thus, GSK3β terminates Arc expression and limits its effect on dendritic spine morphology. Taken together, the results identify GSK3α/β-catalyzed Arc phosphorylation and degradation as a novel mechanism for controlling the duration of Arc expression and function

Concentration and Methylation of Cell-Free DNA from Blood Plasma as Diagnostic Markers of Renal Cancer

The critical point for successful treatment of cancer is diagnosis at early stages of tumor development. Cancer cell-specific methylated DNA has been found in the blood of cancer patients, indicating that cell-free DNA (cfDNA) circulating in the blood is a convenient tumor-associated DNA marker. Therefore methylated cfDNA can be used as a minimally invasive diagnostic marker. We analysed the concentration of plasma cfDNA and methylation of six tumor suppressor genes in samples of 27 patients with renal cancer and 15 healthy donors as controls. The cfDNA concentrations in samples from cancer patients and healthy donors was measured using two different methods, the SYBR Green I fluorescence test and quantitative real-time PCR. Both methods revealed a statistically significant increase of cfDNA concentrations in cancer patients. Hypermethylation on cfDNA was detected for the LRRC3B (74.1%), APC (51.9%), FHIT (55.6%), and RASSF1 (62.9%) genes in patients with renal cancer. Promoter methylation of VHL and ITGA9 genes was not found on cfDNA. Our results confirmed that the cfDNA level and methylation of CpG islands of RASSF1A, FHIT, and APC genes in blood plasma can be used as noninvasive diagnostic markers of cancer

Structure of monomeric full-length ARC sheds light on molecular flexibility, protein interactions, and functional modalities

he activity‐regulated cytoskeleton‐associated protein (ARC) is critical for long‐term synaptic plasticity and memory formation. Acting as a protein interaction hub, ARC regulates diverse signalling events in postsynaptic neurons. A protein interaction site is present in the ARC C‐terminal domain (CTD), a bilobar structure homologous to the retroviral Gag capsid domain. We hypothesized that detailed knowledge of the three‐dimensional molecular structure of monomeric full‐length ARC is crucial to understand its function; therefore, we set out to determine the structure of ARC to understand its various functional modalities. We purified recombinant ARC and analyzed its structure using small‐angle X‐ray scattering and synchrotron radiation circular dichroism spectroscopy. Monomeric full‐length ARC has a compact, closed structure, in which the oppositely charged N‐terminal domain (NTD) and CTD are juxtaposed, and the flexible linker between them is not extended. The modeled structure of ARC is supported by intramolecular live‐cell Förster resonance energy transfer imaging in rat hippocampal slices. Peptides from several postsynaptic proteins, including stargazin, bind to the N‐lobe, but not to the C‐lobe, of the bilobar CTD. This interaction does not induce large‐scale conformational changes in the CTD or flanking unfolded regions. The ARC NTD contains long helices, predicted to form an anti‐parallel coiled coil; binding of ARC to phospholipid membranes requires the NTD. Our data support a role for the ARC NTD in oligomerization as well as lipid membrane binding. The findings have important implications for the structural organization of ARC with respect to distinct functions, such as postsynaptic signal transduction and virus‐like capsid formation