RESUMO
The beginning of the 21st century has seen an increasing number of digital medical devices (DMDs) arrive on the European market, bringing major benefits and changes for society. DMDs are unique in that they bring intelligence to the organisation of care, and generate and collect a wealth of real-life data with ultra-fast life cycles. They have specific requirements, particularly in terms of data security and interoperability. In France and Europe, the construction of evidence, the assessment process and evaluation methodologies with a view to purchase or reimbursement must adjust to these changes, given the specific features of these technologies. This digital leap has opened up new perspectives for healthcare, along with economic, ethical and regulatory issues. The challenge is to assess the clinical and organisational impact, reliability, safety, interoperability, efficiency and budgetary impact of DMDs in line with the requirements of new standards, guidelines and regulations. This should result in a coherent, pragmatic and proportionate evaluation, so that public decision-makers and buyers can take advantage of the potential opportunities that these digital devices offer to improve healthcare delivery. Thus, a fair and informed evaluation of DMDs would emerge, providing a solid basis to steer their inclusion into contemporary medical practices. This fundamental issue of evaluation, linked to the digital nature of these MDs, is what the round table, comprising experts from academia and/or hospitals, institutions and industry, sought to resolve. Discussions led to proposals on how DMDs should be evaluated, bearing in mind their complexity. The round table set out to identify the bottlenecks in the entire evaluation process, from the CE marking phase, compliance with French safety and interoperability requirements, through to national or local evaluation, in order to inform a purchasing policy and draw up proposals covering the entire spectrum. Ten concrete recommendations were put forward by the round table, aimed at improving the evaluation process by making it clearer and more adaptable, thus offering greater flexibility in the evaluation and decision-making stages. This well-thought-out approach is designed to facilitate a comprehensive and flexible evaluation of DMDs given the constantly evolving technological context.
Assuntos
Atenção à Saúde , Humanos , Reprodutibilidade dos Testes , Europa (Continente) , FrançaRESUMO
The different alleles at the (TG)m(T)n polymorphic loci at the 3' end of the human CFTR intron 8 determine the efficiency by which exon 9 is spliced. We identified a novel TG12T3 allele in a congenital bilateral absence of vas deferens (CBAVD) patient who carries a [TG11T7; p.Phe508Cys; p.Met470Val] haplotype on the other chromosome. To better understand the complex regulation of exon 9 splicing, we analyzed the levels of correctly spliced CFTR transcripts in six CFTR-expressing epithelial cell lines derived from lung, colon, testis, vas deferens, and epididymis transiently transfected with four CFTR minigenes (pTG11T7, pTG12T7, pTG12T5, and pTG12T3). In this work, we show that a decrease in the Ts at the polymorphic locus in a TG12 background determines a cell-type dependent reduction in exon 9+ transcripts that is not related to the basal splicing efficiency in the cell line. These data emphasize the role of the T5 allele in CBAVD and identify the T3 allele as a severe cystic fibrosis (CF) disease-causing mutation. Finally, UV cross-linking experiments demonstrated that tissue-specific trans-acting splicing factors do not contribute to the different patterns of exon 9 splicing found between the cell lines. However, we observed that lower numbers of Ts can alter the binding of TDP-43 (TDP43 or TARDBP) to its specific target ug12 in a tissue-specific manner. Our results support the idea that the ratio of general splicing factors plays a role in the tissue variability of exon 9 alternative splicing.
Assuntos
Alelos , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Ducto Deferente/anormalidades , Adulto , Processamento Alternativo , Células CACO-2 , Linhagem Celular , Fibrose Cística/genética , DNA/metabolismo , Proteínas de Ligação a DNA/metabolismo , Epididimo , Éxons , Células HeLa , Humanos , Masculino , Mutação , Oligospermia/genética , Ligação Proteica , Sequências Repetitivas de Ácido Nucleico , Timidina/análise , Transcrição Gênica , TransfecçãoRESUMO
Although Becker muscular dystrophy (BMD; MIM 300376) is mainly caused by gross deletions of the dystrophin gene, the nature of the mutations involved in the remaining cases is of importance because of the milder clinical course of Becker. We have extensively characterized the mRNA changes associated with five novel point mutations giving rise to a Becker phenotype, which confirm that Becker arises largely due to alterations in splicing. In two cases the milder phenotype arises because of exon skipping, leading to an in-frame deletion (c.1603-2A>C and c.4250T>A). In further two cases intronic mutations (c.4519-5C>G and c.961-5925A>C) result in complex splicing changes, but with some residual normal transcripts. The last case, c.10412T>A (p.Leu3471X), results in a truncated transcript missing only part of the COOH terminal of the protein, suggesting that this region is not crucial for dystrophin function. The detection of a low amount of dystrophin in this patient could be attributable to a reduced efficiency of nonsense-mediated decay. The results emphasize that mRNA analysis is important in defining Becker mutations and will be of value in assessing various gene therapy strategies.
Assuntos
Distrofina/genética , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/metabolismo , Mutação , Fenótipo , Adolescente , Adulto , Sequência de Bases , Criança , Códon sem Sentido , Mutação da Fase de Leitura , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Distrofia Muscular de Duchenne/fisiopatologia , Mutagênese Insercional , Deleção de SequênciaRESUMO
BACKGROUND: Although high throughput technologies for gene profiling are reliable tools, sample/tissue heterogeneity limits their outcomes when applied to identify molecular markers. Indeed, inter-sample differences in cell composition contribute to scatter the data, preventing detection of small but relevant changes in gene expression level. To date, attempts to circumvent this difficulty were based on isolation of the different cell structures constituting biological samples. As an alternate approach, we developed a tissue compartment analysis (TCA) method to assess the cell composition of tissue samples, and applied it to standardize data and to identify biomarkers. METHODOLOGY/PRINCIPAL FINDINGS: TCA is based on the comparison of mRNA expression levels of specific markers of the different constitutive structures in pure isolated structures, on the one hand, and in the whole sample on the other. TCA method was here developed with human kidney samples, as an example of highly heterogeneous organ. It was validated by comparison of the data with those obtained by histo-morphometry. TCA demonstrated the extreme variety of composition of kidney samples, with abundance of specific structures varying from 5 to 95% of the whole sample. TCA permitted to accurately standardize gene expression level amongst >100 kidney biopsies, and to identify otherwise imperceptible molecular disease markers. CONCLUSIONS/SIGNIFICANCE: Because TCA does not require specific preparation of sample, it can be applied to all existing tissue or cDNA libraries or to published data sets, inasmuch specific operational compartments markers are available. In human, where the small size of tissue samples collected in clinical practice accounts for high structural diversity, TCA is well suited for the identification of molecular markers of diseases, and the follow up of identified markers in single patients for diagnosis/prognosis and evaluation of therapy efficiency. In laboratory animals, TCA will interestingly be applied to central nervous system where tissue heterogeneity is a limiting factor.