Advanced therapy medicinal products : new strategies for clinical applications of cell and gene therapy

Advanced therapy medicinal products (ATMPs) have a massive potential to address existing unmet medical needs. Specifically, gene therapy medicinal products (GTMPs) may potentially provide cure for several genetic diseases. Despite much research conducted in this field, only a modest number of products are approved and available. This thesis intends to develop an end-to-end understanding of ATMPs, identifying regulatory and patient access hurdles on gene therapy use In Chapter 1, broad research conducted in this field over the last few decades is explored as well as different clinical applications investigated worldwide. These are based on diverse strategies that range from direct gene replacement or addition to more complex pathways such as specific gene editing or RNA targeting. Important safety risks, limited efficacy, manufacturing hurdles, or ethical conflicts may represent challenges in the success of a candidate GTMP. During the development process, it is fundamental to take such aspects into account and establish overcoming strategies. Then, the current European legal framework of ATMPs is reviewed and an overview of the clinical applications for approved and investigational GTMPs is provided. In Europe, the ATMP regulation was fully implemented in 2009 and, at this point, the Committee for Advanced Therapies was created as a dedicated group of specialists to evaluate medicinal products requiring specific expertise in this area. In Chapter 2, major objections, issues, or concerns raised during the Marketing Authorization Application (MAA) for GTMPs between 2009 and 2017 were identified. During the first few years following CAT establishment, quality issues were often identified as major deficiencies, whereas issues at the nonclinical level appeared to be less frequent. Clinical efficacy and safety issues appeared to have a major role in unsuccessful MAA outcome for GTMPs. Most deficiencies were addressed through clarification during the MAA review or in post-marketing settings. The MAA procedure for GTMPs is complex and it is anticipated that continuous MAA submissions will further enhance the experience of both regulators and applicants, reducing the attrition rate for approval. Despite having a positive Marketing Authorization, this does not mean that these products are being used in clinical practice. In Chapter 3, a full set of hurdles potentially preventing patient access to Gene Therapies is identified based on the most recently available literature. A review of the literature using a systematic approach in two distinct databases was performed by identifying relevant, peerreviewed publications, between 2012 and 2018. Seven major topics were identified as potential patient access hurdles, namely affordability, assessment of value, development of therapy, ethical/social factors, evidence generation, operational implementation and regulatory hurdles. From these, twenty-five additional subthemes were further identified. The most frequently mentioned obstacle in the literature is related to the affordability aspect especially focusing on high cost of therapy (84%) and therapy payment/reimbursement (51%). Importantly, the evidence generation focusing on limited trial outcomes (81%) seems to be a strong obstacle in patient access to these therapies. In Chapter 4, a global discussion on the results obtained in chapter 2 and 3 is presented and summarized in the context of the current body of evidence, as well as the current GTMP landscape. A growing number of Gene Therapies are expected to be developed and made available to patients and health care professionals. This thesis contributed to understanding all hurdles, in a complete and integrated fashion, so that strategies are timely established to ensure gene therapy’s benefits are provided to patients and to the society

Differential Expression of miRNAs in Amyotrophic Lateral Sclerosis Patients

Funding Information: Open access funding provided by FCT|FCCN (b-on). This research was funded by Fundação de Ciência e Tecnologia (FCT), grant numbers PTDC/MEC-NEU/31195/2017, UID/BIM/0009/2020 and UIDP/00009/2020. Publisher Copyright: © 2023, The Author(s).Amyotrophic lateral sclerosis (ALS) is a progressive motor neuron disease that affects nerve cells in the brain and spinal cord, causing loss of muscle control, muscle atrophy and in later stages, death. Diagnosis has an average delay of 1 year after symptoms onset, which impairs early management. The identification of a specific disease biomarker could help decrease the diagnostic delay. MicroRNA (miRNA) expression levels have been proposed as ALS biomarkers, and altered function has been reported in ALS pathogenesis. The aim of this study was to assess the differential expression of plasma miRNAs in ALS patients and two control populations (healthy controls and ALS-mimic disorders). For that, 16 samples from each group were pooled, and then 1008 miRNAs were assessed through reverse transcription-quantitative polymerase chain reaction (RT-qPCR). From these, ten candidate miRNAs were selected and validated in 35 ALS patients, 16 ALS-mimic disorders controls and 15 healthy controls. We also assessed the same miRNAs in two different time points of disease progression. Although we were unable to determine a miRNA signature to use as disease or condition marker, we found that miR-7-2-3p, miR-26a-1-3p, miR-224-5p and miR-206 are good study candidates to understand the pathophysiology of ALS.publishersversionepub_ahead_of_prin

Neuropathies - Guillain-Barré syndrome: rehabilitation

Foram revisados artigos nas bases de dados do MedLine (PubMed) e outras fontes de pesquisa, sem limite de tempo. A estratégia de busca utilizada baseou-se em perguntas estruturadas na forma P.I.C.O. (das iniciais "Paciente", "Intervençao", "Controle", "Outcome"). Articles in the MedLine (PubMed) database and other research sources were reviewed, with no age limit. The search strategy used was based on structured questions in the P.I.C.O. format (from the initials: Patient, Intervention, Control and Outcome).&nbsp

Fluorometric determination of ethidium bromide efflux kinetics in Escherichia coli

This is an Open Access article distributed under the terms of the Creative Commons Attribution Licens

Evidences of organic acids exudation in aluminium stress responses of two Madeiran wheat (Triticum aestivum L.) landraces

Two wheat (Triticum aestivum L.) Madeiran landraces were subjected to 100 μM and 200 μM of aluminium (Al) in hydroponic culture, assessing the organic acid exudation role in plant’s responses to this metal. Samples of initial landrace populations (F0), F3 and haplodiploid lines (DH) were evaluated using standard tests: eriochrome cyanine R staining, root elongation and callose accumulation in roots. Root exudates were obtained to determine if the accumulation of malic and citric acids in hydroponic medium was a response to Al exposure. Additionally, the presence of ALMT1 gene was determined using five microsatellite markers. Standard tests confirmed that ISOP 76 was Al tolerant and ISOP 239, Al susceptible. ISOP 76, in the presence of 100 μM Al, exuded substantially more malic acid (12.87 to 43.33 mg/L), than ISOP 239 (3.65 to 7.72 mg/L). The levels of both organic acid exudation were substantially lower in ISOP 239 than in the ISOP 76. In the presence of 200 μM Al, ISOP 76 F0 shows a higher root elongation ratio (better tolerates Al), but the DH line was the one that exuded higher content of malic acid. Different gene alleles and promoters were detected in both landraces. Molecular differences could explain the observed dissimilarity in organic acid exudation response to Al stress.The authors are grateful to the Madeiran farmers providing samples of wheat landraces, and to all staff of CGR ISOPlexis Gene bank and of the Department of Biological Sciences of Alberta University for support and access to laboratory facilities. This work got the funding support from the programs INTERREG III-B, contract 05/MAC/4.1/C15 and Madeira PO 14-20, contract CASBio, refª. M1420-01-0145-FEDER-000011.info:eu-repo/semantics/publishedVersio

Streptococcal bacteraemia in patients submitted to hematopoietic stem cell transplantation : the role of tooth brushing and use of chlorhexidine

A retrospective evaluation of 73 consecutive recipients of hematopoietic stem cell transplantation (HSCT) was conducted to investigated the role of oral care and incidence of streptococcal bacteremia in patients submitted to hematopoietic stem cell transplantation. Patients were retrospectively evaluated and divided into group A (GA=38) and group B (GB=35). During hospitalization patients from GA performed oral hygiene daily with extra soft toothbrush and toothpaste besides performing mouth cleaning with an ethanol-free 0.12% chlorhexidine solution tree times a day. In contrast GB patients performed mouth cleaning with extra soft toothbrush and toothpaste, but no chlorhexidine was used. Using the Chi square test it was observed that all patients from GA presented negative blood culture for alpha-hemolytic Streptococcus viridans and Candida albicans and only 1 patient without oral mucositis from GB presented positive blood cultures for Streptococcus intermedius (p=0.48). The results indicate that methodology used for oral care before the HSCT and the practice of tooth brushing during the period were effective in preventing streptococcal bacteremia. Moreover, our data suggest that the mouth cleaning with chlorhexidine during HSCT may be not mandatory

A influência do exercício físico sobre o cortisol e glicose sanguínea de praticantes de atividade física

Introduction: The secretion of cortisol is directly related to the use of glucose by the body, due to its antagonistic function to glucose, so the importance of the simultaneous evaluation of both, in order to be considered factors associated with muscle protein catabolism and hyperglycemia associated with physical exercise. Objective: To analyze the influence of physical exercise on cortisol and blood glucose. Methodology: The study is a bibliographical review of the integrative type, using the databases Pubmed, Scielo and the search engine Google academic, prioritizing works published between the years 2011 and 2018, totaling 11 studies. Results: It was verified the relationship between the results of the obtained biochemical parameters, allowing to evaluate the physical stress related to the intensity of the physical exercise and, in some cases, to correlate with the carbohydrate supplementation. Final considerations: Through the findings, it is concluded that according to the period of time and hours spent practicing physical exercise there is an adaptation of the body to the secretion of cortisol, where the improvement of performance may be directly related to the use of carbohydrate and that caloric restriction is not a predictor of stress.Introdução: A secreção do cortisol está diretamente relacionada à utilização da glicose pelo organismo, devido a sua função antagônica à glicose, por isso a importância da avaliação simultânea de ambos, afim de que sejam considerados fatores associados com o catabolismo proteico muscular e hiperglicemia associada ao exercício físico. Objetivo: Analisar a influência do exercício físico sobre o cortisol e a glicose sanguínea. Metodologia: O estudo trata-se de uma revisão bibliográfica do tipo integrativa, utilizou-se para a pesquisa as bases de dados: Pubmed, Scielo e o buscador Google acadêmico, priorizando trabalhos publicados entre os anos de 2011 a 2018, totalizando 11 estudos. Resultados: Foi verificado a relação entre os resultados dos parâmetros bioquímicos obtidos, possibilitando avaliar o estresse físico relacionado à intensidade do exercício físico e, em alguns casos, relacionar com a suplementação de carboidratos. Considerações finais: Por meio dos achados, conclui-se que de acordo com o período de tempo e horas dispendidas a prática de exercício físico tem-se uma adaptação do organismo a secreção do cortisol, onde a melhora do desempenho pode estar diretamente relacionada ao uso de carboidrato e que a restrição calórica não é preditora de estresse

Acute encephalic vascular accident: rehabilitation

Este estudo revisou artigos nas bases de dados do MEDLINE (Pub-Med) e demais fontes de pesquisa, sem limite de tempo. Para tanto, adotou-se a estratégia de busca baseada em perguntas estruturadas na forma (P.I.C.O.) das iniciais: "Paciente"; "Intervençao"; "Controle" e "Outcome". Como descritores utilizaram-se: (cerebrovascular disorders OR Stroke) AND acute AND (movement OR physical therapy modalities OR exercise movement techniques); (cerebrovascular disorders OR stroke) AND (Postural Balance OR Musculoskeletal Physiological Phenomena) AND Rehabilitation AND trunk; (cerebrovascular disorders OR Stroke)AND (virtual reality OR user-computer interface OR video games); cerebrovascular disorders AND (home care services, hospital-based); cerebrovascular disorders AND (contractures OR splint OR orthotic devices); cerebrovascular disorders AND acute AND (rehabilitation OR physical therapy disorders or task performance); cerebrovascular disorders AND (physical therapy modalities OR rehabilitation OR exercise therapy) AND (Home Care Services OR self care); (cerebrovascular disorders OR stroke) AND (electric stimulation OR electric stimulation therapy) AND (muscle tonus OR muscle hypotonia OR muscle hypertonia); cerebrovascular disorders AND (restraint physical OR constraint induced movement therapy OR constraint); (cerebrovascular disordersOR stroke) AND (body weight support OR supported treadmill training OR partial weight bearing); (cerebrovascular disorders OR stroke) AND transcranial magnetic stimulation; (cerebrovascular disorders OR stroke) AND bandages. Com esses descritores efetivaram-se cruzamentos de acordo com o tema proposto em cada tópico das perguntas (P.I.C.O.). Analisado esse material, foram selecionados os artigos relativos às perguntas e, por meio do estudo dos mesmos, estabeleceram-se as evidências que fundamentaram às diretrizes do presente documento.This study revised articles from the MEDLINE (PubMed) databases and other research sources, with no time limit. To do so, the search strategy adopted was based on (P.I.C.O.) structured questions (from the initials "Patient"; "Intervention"; "Control" and "Outcome". As keywords were used: (cerebrovascular disorders OR Stroke) AND acute AND (movement OR physical therapy modalities OR exercise movement techniques); (cerebrovascular disorders OR stroke) AND (Postural Balance OR Musculoskel et al. Physiological Phenomena) AND Rehabilitation AND trunk; (cerebrovascular disorders OR Stroke) AND (virtual reality OR user-computer interface OR video games); cerebrovascular disorders AND (home care services, hospital-based); cerebrovascular disorders AND (contractures OR splint OR orthotic devices); cerebrovascular disorders AND acute AND (rehabilitation OR physical therapy disorders or task performance); cerebrovascular disorders AND (physical therapy modalities OR rehabilitation OR exercise therapy) AND (Home Care Services OR self care); (cerebrovascular disorders OR stroke) AND (electric stimulation OR electric stimulation therapy) AND (muscle tonus OR muscle hypotonia OR muscle hypertonia); cerebrovascular disorders AND (restraint physical OR constraint induced movement therapy OR constraint); (cerebrovascular disorders OR stroke) AND (body weight support OR supported treadmill training OR partial weight bearing); (cerebrovascular disorders OR stroke) AND transcranial magnetic stimulation; (cerebrovascular disorders OR stroke) AND bandages. With the above keywords crossings were performed according to the proposed theme in each topic of the (P.I.C.O.) questions. After analyzing this material, articles regarding the questions were selected and, by studying those, the evidences that fundamented the directives of this document were established

Escore preditor da eficácia da quimioterapia pré-operatório em carcinoma de mama localmente avançado / Predictive score of preoperative chemotherapy effectiveness in locally advanced breast carcinoma

Com intuito de desenvolver um escore preditor da resposta quimioterápica neoadjuvante, propusemos a cintilografia de mamas com sestamibi-99mTc (CM), associada a fatores prognósticos como método preditor. Foram estudadas 65 pacientes com diagnóstico de carcinoma ductal submetidas à cintilografia de mamas pré-quimioterapia neoadjuvante (QT neo), painel imunohistoquímico e avaliação do espécime cirúrgico. Foram geradas áreas de interesse com contagens/pixels sobre a lesão e na mama contralateral e criados índices com a razão dessas contagens. Foi construído um modelo de regressão logística utilizando como variáveis, idade, índice, localização do tumor, painel imunohistoquímico e comprometimento axilar, obtendo-se valores preditos que em função da resposta patológica completa foi criada uma curva ROC, para que se obtivesse escore de maior sensibilidade e especificidade. A área da curva ROC foi significativa e o escore obtido foi de 0,728 com sensibilidade, especificidade e valores preditivos positivo e negativo de 81,6%, 81,25%, 93% e 59%, respectivamente. O escore preditor multifatorial é um método eficaz na predição da resposta quimioterápica neoadjuvante, uma vez, que reflete a biologia do tumor frente ao esquema quimioterápico, identificando precocemente as pacientes que não se beneficiariam deste tratamento

Construção de um referencial de avaliação de desempenho do estudante de línguas em estágio pedagógico: problemas e desafios

Este artigo tem como objetivo apresentar os mecanismos de construção e as questões teóricas e metodológicas subjacentes ao processo de elaboração de um referencial de avaliação do desempenho de professores de línguas em formação inicial nos cursos de mestrado em ensino e de português língua segunda/língua estrangeira da Faculdade de Letras da Universidade do Porto (FLUP) tornando visíveis algumas das opções teóricas e metodológicas assumidas. O referencial de avaliação de desempenho do estudante em estágio pedagógico constitui-se como documento revelador da avaliação da formação de professores de línguas da FLUP e configura o perfil do futuro professor de línguas. Em 2015, um grupo constituído por docentes e investigadores da FLUP, em colaboração com docentes orientadores de estágio, iniciou um processo de elaboração de um referencial de avaliação no âmbito da formação de professores de línguas. Esse processo desenvolveu-se em diversas etapas, tendo havido monitorização do documento por parte de um grupo de orientadores cooperantes, de supervisores e de outros professores de alguma forma associados ao estágio pedagógico. Neste artigo, encontra-se a descrição das dificuldades e desafios relativos a cada etapa e a explicitação das referências teóricas em que se fundam as opções assumidas.The purpose of this paper is to describe the process undertaken (incorporating both theoretical and methodological perspectives) in the construction of a unified set of descriptors intended for the assessment of student teachers of pre-service master degrees in the teaching of Portuguese as a foreign language and other modern languages at the Faculty of Arts and Humanities of the University of Porto (FLUP). This assessment document covers the proficiency and performance of student-teachers during their practicum and delimits the professional profile of future teachers of modern languages. In 2015, a group consisting of teachers and researchers from FLUP, in collaboration with school mentors, started the process of constructing a unified assessment document for use in the training of teachers of modern languages. This process developed through various stages, including the piloting of the document by several school mentors, FLUP supervisors and other faculty staff involved in initial teacher education. This article highlights the difficulties and challenges encountered at each stage in the development of this document; in addition, explicit reference is made to the theoretical foundations supporting the decisions made