Browsing by Author "Nóbrega, Clévio"
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- Advanced therapy medicinal products development - from guidelines to medicines in the marketPublication . Frederico, Catarina; Vieira da Conceição, André Filipe; Nóbrega, Clévio; Mendonça, Liliana S.In Europe, Advanced Therapy Medicinal Products (ATMPs) include medicines based on gene therapy, somaticcell therapy, tissue-engineered products, and combined ATMPs. ATMPs constitute an emerging and innovative class of medicines used to treat multiple pathologies and are particularly relevant in pathologies where therapeutic options are limited and require high medical needs. These therapies act, among others, through the insertion of recombinant nucleic acids, including genes, to promote a therapeutic effect and through the restoration of cell functions, and repairing or replacing damaged cells and tissues impaired in pathological conditions. Despite their unique potential, these therapies face challenges related to scientific complexity, production processes, regulatory approval, and market access that hinder their development and availability. Based on official European guidelines, the present review explores the current regulatory framework for the non-clinical and clinical development of advanced therapies. We aimed to discuss the regulations applied to the different types of ATMPs, as well as the challenges associated with their development until these therapies reach the market. Accordingly, topics such as the implementation of proof-of-concept studies to provide evidence supporting the potential clinical effect; biodistribution studies to evaluate tissue distribution and persistence; and toxicology studies to assess potential undesirable effects, integration potential of viral vectors, tumorigenicity, and germline transmission, are discussed. This work also covers some of the ATMPs available to patients on the EU market.
- Autophagy in Spinocerebellar ataxia type 2, a dysregulated pathway, and a target for therapyPublication . Marcelo, Adriana; Afonso, Inês T.; Afonso-Reis, Ricardo; Brito, David V. C.; Costa, Rafael G.; Rosa, Ana; Alves-Cruzeiro, João; Ferreira, Benedita; Henriques, Carina; Nobre, Rui J.; Matos, Carlos A; de Almeida, Luís Pereira; Nóbrega, ClévioSpinocerebellar ataxia type 2 (SCA2) is an incurable and genetic neurodegenerative disorder. The disease is characterized by progressive degeneration of several brain regions, resulting in severe motor and non-motor clinical manifestations. The mutation causing SCA2 disease is an abnormal expansion of CAG trinucleotide repeats in the ATXN2 gene, leading to a toxic expanded polyglutamine segment in the translated ataxin-2 protein. While the genetic cause is well established, the exact mechanisms behind neuronal death induced by mutant ataxin-2 are not yet completely understood. Thus, the goal of this study is to investigate the role of autophagy in SCA2 pathogenesis and investigate its suitability as a target for therapeutic intervention. For that, we developed and characterized a new striatal lentiviral mouse model that resembled several europathological hallmarks observed in SCA2 disease, including formation of aggregates, neuronal marker loss, cell death and neuroinflammation. In this new model, we analyzed autophagic markers, which were also analyzed in a SCA2 cellular model and in human post-mortem brain samples. Our results showed altered levels of SQSTM1 and LC3B in cells and tissues expressing mutant ataxin-2. Moreover, an abnormal accumulation of these markers was detected in SCA2 patients’ striatum and cerebellum. Importantly, the molecular activation of autophagy, using the compound cordycepin, mitigated the phenotypic alterations observed in disease models. Overall, our study suggests an important role for autophagy in the context of SCA2 pathology, proposing that targeting this pathway could be a potential target to treat SCA2 patients.
- Autophagy in Spinocerebellar Ataxia Type 3: From pathogenesis to therapeuticsPublication . Paulino, Rodrigo; Nóbrega, ClévioMachado–Joseph disease (MJD) or spinocerebellar ataxia 3 (SCA3) is a rare, inherited, monogenic, neurodegenerative disease, and the most common SCA worldwide. MJD/SCA3 causative mutation is an abnormal expansion of the triplet CAG at exon 10 within the ATXN3 gene. The gene encodes for ataxin-3, which is a deubiquitinating protein that is also involved in transcriptional regulation. In normal conditions, the ataxin-3 protein polyglutamine stretch has between 13 and 49 glutamines. However, in MJD/SCA3 patients, the size of the stretch increases from 55 to 87, contributing to abnormal protein conformation, insolubility, and aggregation. The formation of aggregates, which is a hallmark of MJD/SCA3, compromises different cell pathways, leading to an impairment of cell clearance mechanisms, such as autophagy. MJD/SCA3 patients display several signals and symptoms in which the most prominent is ataxia. Neuropathologically, the regions most affected are the cerebellum and the pons. Currently, there are no disease-modifying therapies, and patients rely only on supportive and symptomatic treatments. Due to these facts, there is a huge research effort to develop therapeutic strategies for this incurable disease. This review aims to bring together current state-of-the-art strategies regarding the autophagy pathway in MJD/SCA3, focusing on evidence for its impairment in the disease context and, importantly, its targeting for the development of pharmacological and gene-based therapies.
- Brain-targeted drug delivery - nanovesicles directed to specific brain cells by brain-targeting ligandsPublication . Moreira, Ricardo; Nóbrega, Clévio; Almeida, Luís Pereira de; Mendonça, LilianaNeurodegenerative diseases are characterized by extensive loss of function or death of brain cells, hampering the life quality of patients. Brain-targeted drug delivery is challenging, with a low success rate this far. Therefore, the application of targeting ligands in drug vehicles, such as lipid-based and polymeric nanoparticles, holds the promise to overcome the blood-brain barrier (BBB) and direct therapies to the brain, in addition to protect their cargo from degradation and metabolization. In this review, we discuss the barriers to brain delivery and the different types of brain-targeting ligands currently in use in brain-targeted nanoparticles, such as peptides, proteins, aptamers, small molecules, and antibodies. Moreover, we present a detailed review of the different targeting ligands used to direct nanoparticles to specific brain cells, like neurons (C4-3 aptamer, neurotensin, Tet-1, RVG, and IKRG peptides), astrocytes (Aquaporin-4, D4, and Bradykinin B2 antibodies), oligodendrocytes (NG-2 antibody and the biotinylated DNA aptamer conjugated to a streptavidin core Myaptavin-3064), microglia (CD11b antibody), neural stem cells (QTRFLLH, VPTQSSG, and NFL-TBS.40–63 peptides), and to endothelial cells of the BBB (transferrin and insulin proteins, and choline). Reports demonstrated enhanced brain-targeted delivery with improved transport to the specific cell type targeted with the conjugation of these ligands to nanoparticles. Hence, this strategy allows the implementation of high-precision medicine, with reduced side effects or unwanted therapy clearance from the body. Nevertheless, the accumulation of some of these nanoparticles in peripheral organs has been reported indicating that there are still factors to be improved to achieve higher levels of brain targeting. This review is a collection of studies exploring targeting ligands for the delivery of nanoparticles to the brain and we highlight the advantages and limitations of this type of approach in precision therapies.
- Cordycepin activates autophagy through AMPK phosphorylation to reduce abnormalities in Machado-Joseph disease modelsPublication . Marcelo, Adriana; Brito, Filipa; Carmo-Silva, Sara; Matos, Carlos A.; Alves-Cruzeiro, Joao; Vasconcelos-Ferreira, Ana; Koppenol, Rebekah; Mendonca, Liliana; de Almeida, Luis Pereira; Nóbrega, ClévioMachado-Joseph disease (MJD) is a neurodegenerative disorder caused by an abnormal expansion of citosine-adenine-guanine trinucleotide repeats in the disease-causing gene. This mutation leads to an abnormal polyglutamine tract in the protein ataxin-3 (Atx3), resulting in formation of mutant Atx3 aggregates. Despite several attempts to develop a therapeutic option for MJD, currently there are no available therapies capable of delaying or stopping disease progression. Recently, our group reported that reducing the expression levels of mutant Atx3 lead to a mitigation of several MJD-related behavior and neuropathological abnormalities. Aiming a more rapid translation to the human clinics, in this study we investigate a pharmacological inhibitor of translation-cordycepin-in several preclinical models. We found that cordycepin treatment significantly reduced (i) the levels of mutant Atx3, (ii) the neuropathological abnormalities in a lentiviral mouse model, (iii) the motor and neuropathological deficits in a transgenic mouse model and (iv) the number of ubiquitin aggregates in a human neural model. We hypothesize that the effect of cordycepin is mediated by the increase of phosphorylated adenosine monophosphate-activated protein kinase (AMPK) levels, which is accompanied by a reduction in the global translation levels and by a significant activation of the autophagy pathway. Overall, this study suggests that cordycepin might constitute an effective and safe therapeutic approach for MJD, and probably for the other polyglutamine diseases.
- Current status of gene therapy research in polyglutamine spinocerebellar ataxiasPublication . Afonso-Reis, Ricardo; Afonso, Inês T.; Nóbrega, ClévioPolyglutamine spinocerebellar ataxias (PolyQ SCAs) are a group of 6 rare autosomal dominant diseases, which arise from an abnormal CAG repeat expansion in the coding region of their causative gene. These neurodegenerative ataxic disorders are characterized by progressive cerebellar degeneration, which translates into progressive ataxia, the main clinical feature, often accompanied by oculomotor deficits and dysarthria. Currently, PolyQ SCAs treatment is limited only to symptomatic mitigation, and no therapy is available to stop or delay the disease progression, which culminates with death. Over the last years, many promising gene therapy approaches were investigated in preclinical studies and could lead to a future treatment to stop or delay the disease development. Here, we summed up the most promising of these therapies, categorizing them in gene augmentation therapy, gene silencing strategies, and gene edition approaches. While several of the reviewed strategies are promising, there is still a gap from the preclinical results obtained and their translation to clinical studies. However, there is an increase in the number of approved gene therapies, as well as a constant development in their safety and efficacy profiles. Thus, it is expected that in a near future some of the promising strategies reviewed here could be tested in a clinical setting and if successful provide hope for SCAs patients.
- CYP46A1- gene therapy improves Machado-Joseph disease in mouse modelsPublication . Nóbrega, Clévio; Mendonca, L. S.; Marcelo, Adriana; Lamaziere, A.; Tome, S.; Despres, G.; Matos, C.; Mechmet, Fatich; Langui, D.; den Dunnen, W.; de Almeida, L. P.; Cartier, N.; Alves, S.Aims/Context: Machado-Joseph Disease (MJD) is a neurodegenerative disease associated with extensive neuronal death. Defects in brain cholesterol metabolism may contribute to neurodegenerative diseases.
- Editorial: Advanced (gene and cell) therapies for central nervous system applicationsPublication . Mendonça, Liliana; Webster, Christopher; Boltze, Johannes; Nóbrega, ClévioAdvanced therapies as defined by the European Medicines Agency (EMA) comprises strategies involving gene therapy, cell therapy, and tissue engineering. Overall, these strategies offer a wide range of possibilities to treat and cure diseases, including those affecting the central nervous system (CNS). In this line, Advanced (Gene and Cell) Therapies for Central Nervous System Applications Research Topic was intended to provide a platform for researchers to publish their findings, contributing to the continuous advance of this research area. Nine papers were accepted and published in this Research Topic, from which four described original research data, two papers were reviews and two mini reviews, and one paper focused on hypothesis and theory. In the paper Administration of Variants AAV-PHP.B and AAV-PHP.eB on Brain Transduction in Adult Rhesus Macaques (Arotcarena et al.), presented new data on the biodistribution and CNS transduction efficiencies after lumbar intrathecal bolus delivery of identical doses of either AAV-PH.B:CAG-EGFP or AAV-PHP.eB:CAG-EGFP in rhesus macaque monkeys.
- Editorial: Global excellence in gene and cell therapy: EuropePublication . Mendonça, Liliana; Boltze, Johannes; Nóbrega, ClévioGene therapy changes protein expression in cells through the insertion of genes and/or gene expression-modulating tools, including gene editing. It can be used in the treatment of a variety of diseases, including genetic disorders, by reducing the levels of disease-causing proteins or by increasing the expression of missing or protective proteins. Cell therapy by the transplantation of cells or tissues aims to repair, regenerate, or replace a specific cell population and/or tissue to reestablish the physiological function performed by the cells/tissues to replace. In this line, Global Excellence in the Gene and Cell Therapy: Europe Research Topic was launched with the goal of highlighting the latest advancements in Gene and Cell Therapy field and reflecting on the future challenges faced by researchers across Europe. Six papers were accepted and published for this Research Topic, from which three describe original research data, one is a systematic review, one is a narrative paper, and the last one is a perspective article.
- Establishment of an induced pluripotent cell line (ABCRIi001-A) from an elderly female for ageing researchPublication . Esteves, Filipa; Vilhena Catarino Brito, David; Rajado, Ana Teresa; Silva, Nádia; Apolónio, Joana; Roberto, Vânia; Andrade, Raquel; Calado, Sofia; Faleiro, Maria Leonor; Albuquerque Andrade de Matos, Carlos Adriano; Marques, Nuno; Marreiros, Ana; Nzwalo, Hipólito; Pais, Sandra; Palmeirim, Isabel; Simãoa, Sónia; Joaquim, Natércia; Miranda, Rui; Pêgas, António; Raposo, Daniela Marques; Sardo, Ana; Araújo, Inês; Nóbrega, Clévio; Castelo-Branco, Pedro; Bragança, JoséHuman induced pluripotent stem cells (hiPSCs) hold promises to model and understand human diseases, including those associated with ageing. Here, we describe ABCRIi001-A, a hiPSC line generated from peripheral blood mononuclear cells (PBMCs) of a 79-year-old female enrolled in a study for development of an ageing score (ALFA Score). PBMCs were reprogrammed using three Sendai virus-based reprogramming vectors (hKOS, hc-Myc, and hKlf4). ABCRIi001-A showed normal morphology and karyotype, viral clearance, absence of genomic aberrations, and their pluripotency was confirmed by expression of pluripotency-related markers and their ability to differentiate into the three germ layers. ABCRIi001-A is valuable for ageing-related studies.