83 research outputs found

    Development of aptamer nanoparticles for treatment of retinal diseases

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    At more advanced states, retinal diseases such as diabetic retinopathy, age-related macular degeneration, and retinal vessel occlusions are characterized by neovascularization, which is usually triggered by a hypoxia episode leading to the overexpression of vascular endothelial growth factor (VEGF) and nucleolin (NCL). The treatments available are mainly corticosteroids or/and antibodies against some angiogenic molecules. Regarding the low efficacy of these therapies and their side effects, aptamers are an emerging tool that can target these proteins with high specificity, such as antibodies. Furthermore, aptamers are easier to synthesize and present higher stability and lower immunogenicity. AT11-L0 is an aptamer derivative of AS1411 composed of G-rich sequences which can adopt G-quadruplex (G4) structure and target NCL, a protein that can act as a co-receptor for several growth factors. Hence, this study aimed to characterize the AT11-L0 structure and its interaction with several ligands (stabilizing molecules or drugs) for NCL targeting. The AT11-L0-drug/molecule complex was then incorporated in a nanoparticle to increase the bioavailability of the aptamer-based drug in the formulation. To achieve these objectives, we have performed biophysical studies, such as nuclear magnetic resonance, circular dichroism, and fluorescence experiments. Following biophysical characterization studies, we synthesised gold nanoparticles and liposomes containing AT11-L0 aptamer-drug complex. Finally, liposomes and the encapsulated drugs were tested on Human umbilical vein endothelial cells (HUVEC) model to assess their antiangiogenic capacity. The results showed that AT11-L0 aptamer-drug complex has a high stability with melting temperatures reaching 45 ºC to 60 ºC allowing an efficient targeting of NCL with a KD in the order of micromolar. HUVEC antiangiogenic assay showed that, although liposomes did not promote an increase in the antiangiogenic effect of the tested drugs, C8 showed an antiangiogenic effect that was not previously described, at lower concentration. This result can be a basis for the development of a new therapeutic approach using C8 and G4 aptamers targeting NCL.Em estágios mais avançados, as doenças da retina como a retinopatia diabética, degeneração macular associada à idade e oclusões da retina são caracterizadas por neovascularização, que pode ser desencadeada por um episódio de hipoxia que envolve a sobre expressão do fator de crescimento endotelial vascular (VEGF) e da nucleolina (NCL). Os tratamentos disponíveis são principalmente corticosteroides e/ou anticorpos específicos para algumas moléculas angiogénicas. Para ultrapassar a baixa eficácia destas terapias e os seus efeitos secundários, os aptameros surgiram como uma ferramenta emergente que, de modo semelhante aos anticorpos, visam estas proteínas com elevada especificidade. Além disso, os aptameros são mais fáceis de sintetizar e apresentam maior estabilidade e menor imunogenicidade. O AT11-L0 é um derivado do aptamero AS1411, composto por sequências ricas em G, que pode adotar estruturas em G-quadruplex e reconhecer a NCL, uma proteína que pode atuar como coreceptora de vários fatores de crescimento. Assim, este estudo visou caracterizar a estrutura do AT11-L0 e a sua interação com vários ligandos (moléculas estabilizadoras ou fármacos), para ligar de modo específico à NCL. Este complexo foi posteriormente incorporado num nanossistema, para aumentar a biodisponibilidade do fármaco e do aptamero usados na formulação. Para alcançar estes objetivos, foram realizados estudos biofísicos, tais como ressonância magnética nuclear, dicroísmo circular e experiências de fluorescência. Após estudos de caracterização biofísica, foram sintetizadas nanopartículas de ouro e lipossomas contendo o complexo aptamero-fármaco. Finalmente, os lipossomas e os fármacos foram testados num modelo de células endoteliais de veia umbilical humana (HUVEC) para analisar a sua capacidade antiangiogénica. Os resultados mostraram que os complexos aptamero-fármaco têm uma elevada estabilidade térmica, com temperaturas de fusão que atingem 45 ºC a 60 ºC, permitindo uma interação com a NCL com um KD na ordem dos micromolares. O ensaio antiangiogénico mostrou que, embora os lipossomas não promovam um aumento do efeito dos fármacos testados, o C8 apresentou um efeito antiangiogénico que nunca foi descrito, mesmo quando usado em baixa concentração. Este resultado pode permitir uma nova abordagem utilizando o C8 associado a aptameros de G4 que visem a NCL

    Artificial intelligence in sickle disease

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    Artificial intelligence (AI) is rapidly becoming an established arm in medical sciences and clinical practice in numerous medical fields. Its implications have been rising and are being widely used in research, diagnostics, and treatment options for many pathologies, including sickle cell disease (SCD). AI has started new ways to improve risk stratification and diagnosing SCD complications early, allowing rapid intervention and reallocation of resources to high-risk patients. We reviewed the literature for established and new AI applications that may enhance management of SCD through advancements in diagnosing SCD and its complications, risk stratification, and the effect of AI in establishing an individualized approach in managing SCD patients in the future. Aim: to review the benefits and drawbacks of resources utilizing AI in clinical practice for improving the management for SCD cases.Open Access funding provided by the Qatar National Library.Scopu

    The Retina in Health and Disease

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    Vision is the most important sense in higher mammals. The retina is the first step in visual processing and the window to the brain. It is not surprising that problems arising in the retina lead to moderate to severe visual impairments. We offer here a collection of reviews as well as original papers dealing with various aspects of retinal function as well as dysfunction. New approaches in retinal research are described, such as the expression and localization of the endocannabinoid system in the normal retina and the role of cannabinoid receptors that could offer new avenues of research in the development of potential treatments for retinal diseases. Moreover, new insights are offered in advancing knowledge towards the prevention and cure of visual pathologies, mainly AMD, RP, and diabetic retinopathy

    In Vitro Models for the Evaluation of Antisense Oligonucleotides in Skin

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    The genodermatosis dystrophic epidermolysis bullosa (DEB) is caused by mutations in the COL7A1 gene which encodes type VII collagen (C7). In the cutaneous basement membrane zone, C7 secures attachment of the epidermal basal keratinocyte to the papillary dermis by means of anchoring fibril formation. The complete absence of these anchoring fibrils leads to severe blistering of skin and mucosa upon the slightest friction and early mortality. To date, although preclinical advances toward therapy are promising, treatment for the disease is merely symptomatic. Therefore, research into novel therapeutics is warranted.Antisense oligonucleotide (ASO)-mediated exon skipping is such a therapy . Clinical examination of naturally occurring exon skipping suggested that this mechanism could most likely benefit the most severely affected patients. The severe form of DEB is caused by biallelic null mutations. Exon skipping aims to bind an ASO to the mutated exon of the pre-mRNA in the cell nucleus. Thereby, the ASO inhibits the recognition of the mutated exon by the splicing machinery, and as a result, the mutated exon is spliced out from the mRNA with its surrounding introns, i.e., it is skipped. Here, we describe in vitro methods to evaluate ASO-mediated exon skipping in a preclinical setting

    Evaluation of PD-L1 expression in various formalin-fixed paraffin embedded tumour tissue samples using SP263, SP142 and QR1 antibody clones

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    Background & objectives: Cancer cells can avoid immune destruction through the inhibitory ligand PD-L1. PD-1 is a surface cell receptor, part of the immunoglobulin family. Its ligand PD-L1 is expressed by tumour cells and stromal tumour infltrating lymphocytes (TIL). Methods: Forty-four cancer cases were included in this study (24 triple-negative breast cancers (TNBC), 10 non-small cell lung cancer (NSCLC) and 10 malignant melanoma cases). Three clones of monoclonal primary antibodies were compared: QR1 (Quartett), SP 142 and SP263 (Ventana). For visualization, ultraView Universal DAB Detection Kit from Ventana was used on an automated platform for immunohistochemical staining Ventana BenchMark GX. Results: Comparing the sensitivity of two different clones on same tissue samples from TNBC, we found that the QR1 clone gave higher percentage of positive cells than clone SP142, but there was no statistically significant difference. Comparing the sensitivity of two different clones on same tissue samples from malignant melanoma, the SP263 clone gave higher percentage of positive cells than the QR1 clone, but again the difference was not statistically significant. Comparing the sensitivity of two different clones on same tissue samples from NSCLC, we found higher percentage of positive cells using the QR1 clone in comparison with the SP142 clone, but once again, the difference was not statistically significant. Conclusion: The three different antibody clones from two manufacturers Ventana and Quartett, gave comparable results with no statistically significant difference in staining intensity/ percentage of positive tumour and/or immune cells. Therefore, different PD-L1 clones from different manufacturers can potentially be used to evaluate the PD- L1 status in different tumour tissues. Due to the serious implications of the PD-L1 analysis in further treatment decisions for cancer patients, every antibody clone, staining protocol and evaluation process should be carefully and meticulously validated

    Advance Nanomaterials for Biosensors

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    The book provides a comprehensive overview of nanostructures and methods used to design biosensors, as well as applications for these biosensor nanotechnologies in the biological, chemical, and environmental monitoring fields. Biological sensing has proven to be an essential tool for understanding living systems, but it also has practical applications in medicine, drug discovery, food safety, environmental monitoring, defense, personal security, etc. In healthcare, advancements in telecommunications, expert systems, and distributed diagnostics are challenging current delivery models, while robust industrial sensors enable new approaches to research and development. Experts from around the world have written five articles on topics including:Diagnosing and treating intraocular cancers such as retinoblastoma; Nanomedicine in cancer management; Engineered nanomaterials in osteosarcoma diagnosis and treatment; Practical design of nanoscale devices; Detect alkaline phosphatase quantitatively in clinical diagnosis; Progress in the area of non-enzymatic sensing of dual/multi biomolecules; Developments in non-enzymatic glucose and H2O2 (NEGH) sensing; Multi-functionalized nanocarrier therapies for targeting retinoblastoma; Galactose functionalized nanocarriers; Sensing performance, electro-catalytic mechanism, and morphology and design of electrode materials; Biosensors along with their applications and the benefits of machine learning; Innovative approaches to improve the NEGH sensitivity, selectivity, and stability in real-time applications; Challenges and solutions in the field of biosensors

    In Vivo Models for the Evaluation of Antisense Oligonucleotides in Skin

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    Here, we describe an in vivo model in which antisense oligonucleotides were preclinically evaluated in reconstituted patient and healthy control skin. The aim was to investigate the effect of antisense oligonucleotides upon local or systemic administration. This allows for clinically relevant evaluation of antisense oligonucleotides in an in vivo setting. In this model, primary human keratinocytes and fibroblasts were placed into silicone grafting chambers, implanted onto the back of athymic nude mice. After sufficient cells were expanded, within a few weeks, human skin grafts were generated with a high success rate. These mice bearing grafts were subsequently treated with antisense oligonucleotides targeting exon 105 of the COL7A1 gene which encodes type VII collagen. Patients completely lacking expression of type VII collagen develop severe blistering of skin and mucosa, i.e., recessive dystrophic epidermolysis bullosa. In this chapter, we describe the in vivo model used for the preclinical evaluation of antisense oligonucleotides as therapeutic approach for recessive dystrophic epidermolysis bullosa

    ABC Transporters in Human Diseases

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    Mammalian ATP-binding cassette (ABC) transporters constitute a superfamily of proteins involved in many essential cellular processes. Most of these transporters are transmembrane proteins and allow the active transport of solutes, small molecules, and lipids across biological membranes. On the one hand, some of these transporters are involved in drug resistance (also referred to as MDR or multidrug resistance), a process known to be a major brake in most anticancer treatments, and the medical challenge is thus to specifically inhibit their function. On the other hand, molecular defects in some of these ABC transporters are correlated with several rare human diseases, the most well-documented of which being cystic fibrosis, which is caused by genetic variations in ABCC7/CFTR (cystic fibrosis transmembrane conductance regulator). In the latter case, the goal is to rescue the function of the deficient transporters using various means, such as targeted pharmacotherapies and cell or gene therapy. The aim of this Special Issue, “ABC Transporters in Human Diseases”, is to present, through original articles and reviews, the state-of-the-art of our current knowledge about the role of ABC transporters in human diseases and the proposed therapeutic options based on studies ranging from cell and animal models to patients

    Drug development progress in duchenne muscular dystrophy

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    Duchenne muscular dystrophy (DMD) is a severe, progressive, and incurable X-linked disorder caused by mutations in the dystrophin gene. Patients with DMD have an absence of functional dystrophin protein, which results in chronic damage of muscle fibers during contraction, thus leading to deterioration of muscle quality and loss of muscle mass over time. Although there is currently no cure for DMD, improvements in treatment care and management could delay disease progression and improve quality of life, thereby prolonging life expectancy for these patients. Furthermore, active research efforts are ongoing to develop therapeutic strategies that target dystrophin deficiency, such as gene replacement therapies, exon skipping, and readthrough therapy, as well as strategies that target secondary pathology of DMD, such as novel anti-inflammatory compounds, myostatin inhibitors, and cardioprotective compounds. Furthermore, longitudinal modeling approaches have been used to characterize the progression of MRI and functional endpoints for predictive purposes to inform Go/No Go decisions in drug development. This review showcases approved drugs or drug candidates along their development paths and also provides information on primary endpoints and enrollment size of Ph2/3 and Ph3 trials in the DMD space
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