Search Results (8)

Host–pathogen interactions and the design of vaccines for aquaculture fish viruses are challenging and call for innovative approaches. This study explores the potential of adenoviral (Ad) vectors Ad5 and chimeric Ad5/40 as gene delivery tools for fish brain cells susceptible to neurotropic viruses. For this purpose, European sea bass (Dicentrarchus labrax) DLB-1 and gilthead seabream (Sparus aurata) SaB-1 brain cell lines were infected with Ad5 or Ad5/40 vectors expressing GFP, and we evaluated their capacity for infection by fluorescence microscopy and flow cytometry, as well as their antiviral innate immune response by the transcription of gene markers (irf3 and mx). We found that both vectors are able to infect DLB-1 and SaB-1 brain cell lines to similar levels, as demonstrated by fluorescence microscopy and flow cytometry, though the infection efficiency was low. In addition, infection with Ad vectors regulated the transcription of genes related to the interferon-mediated antiviral immune response. Our results indicate that the Ad5/40 vector achieves better infection and consistent cellular distribution. These findings suggest that these vectors may offer targeted gene delivery and local immune responses. Full article

Background: The biosafety of gene therapy products remains a major challenge to their introduction into the clinic. In particular, the problem of immunogenicity of viral vectors is the focus of attention. Large animals such as pigs, whose anatomical and physiological characteristics are similar to those of humans, have an advantage in testing vector systems. Methods: We performed a comprehensive in vitro and in vivo study to evaluate the biosafety of a chimeric adenoviral vector carrying a green fluorescent protein gene (Ad5/35F-GFP) in a mini-pig model. Results: Transcriptome and secretome analyses of mini-pig leucocytes transduced with Ad5/35F-GFP revealed changes restraining pro-inflammatory processes and cytokine production. No adverse effects were revealed through the clinical, instrumental, laboratory, and histological examinations conducted within a week after the direct or autologous leucocyte-mediated administration of Ad5/35F-GFP to mini-pigs. The decrease in cytokine levels in the blood of experimental animals is also consistent with the in vitro data and confirms the immune tolerance of mini-pigs to Ad5/35F-GFP. Conclusions: Here, we show the safety of Ad5/35F in a mini-pig model and provide evidence that Ad5/35F is a promising vector for gene therapy. These results advance our understanding of vector–host interactions and offer a solid foundation for the clinical application of this vector. Full article

The natural limitations of regeneration in the CNS are major problems for the treatment of neurological disorders, including ischaemic brain strokes. Among the approaches being actively developed to inhibit post-ischaemic negative consequences is the delivery of therapeutic genes encoding neuroprotective molecules to the brain. Unfortunately, there are currently no proven and available medicines that contain recombinant human genes for the treatment of ischaemic cerebral stroke. Of particular interest is the development of treatments for patients at risk of ischaemic stroke. In the present study, we propose a proof of concept for the use of an autologous, genetically enriched leucoconcentrate temporally secreting recombinant vascular endothelial growth factor (VEGF), glial-cell-line-derived neurotrophic factor (GDNF) and the neural cell adhesion molecule (NCAM) for the treatment of stroke. In a mini-pig ischaemic stroke model, genetically enriched leucoconcentrate was infused 4 h after surgery (gene therapy in acute phase) or 2 days before stroke modelling (preventive gene therapy). On day 21, after the stroke modelling, the post-ischaemic brain recovery was examined by morphologic and immunofluorescence analysis. The benefits of treating a stroke with genetically enriched leucoconcentrate both for preventive purposes and in the acute phase were confirmed by an improved performance in behavioural tests, higher preservation of brain tissue and positive post-ischaemic brain remodelling in the peri-infarct area. These results suggest that the employment of autologous leucocytes enabling the temporary production of the recombinant therapeutic molecules to correct the pathological process in the CNS may be one of the breakthrough approaches in gene therapy. Full article

The contemporary strategy for spinal cord injury (SCI) therapy aims to combine multiple approaches to control pathogenic mechanisms of neurodegeneration and stimulate neuroregeneration. In this study, a novel regenerative approach using an autologous leucoconcentrate enriched with transgenes encoding vascular endothelial growth factor (VEGF), glial cell line-derived neurotrophic factor (GDNF), and neural cell adhesion molecule (NCAM) combined with supra- and sub-lesional epidural electrical stimulation (EES) was tested on mini-pigs similar in morpho-physiological scale to humans. The complex analysis of the spinal cord recovery after a moderate contusion injury in treated mini-pigs compared to control animals revealed: better performance in behavioural and joint kinematics, restoration of electromyography characteristics, and improvement in selected immunohistology features related to cell survivability, synaptic protein expression, and glial reorganization above and below the injury. These results for the first time demonstrate the positive effect of intravenous infusion of autologous genetically-enriched leucoconcentrate producing recombinant molecules stimulating neuroregeneration combined with neuromodulation by translesional multisite EES on the restoration of the post-traumatic spinal cord in mini-pigs and suggest the high translational potential of this novel regenerative therapy for SCI patients. Full article

Gene therapy vectors derived from different viral species have become a fixture in biomedicine, both for direct therapeutic intervention and as tools to facilitate cell-based therapies, such as chimeric antigen receptor-based immunotherapies. On the contrary, extracellular vesicles have only recently gained a massive increase in interest and, concomitantly, knowledge in the field has drastically risen. Viral infections and extracellular vesicle biology overlap in many ways, both with pro- and antiviral outcomes. In this review, we take a closer look at these interactions for the most prominent groups of viral vectors (Adenoviral, Adeno-associated and Retro/Lentiviral vectors) and the possible implications of these overlaps for viral vector technology and its biomedical applications. Full article

Adenoviral viral vectors have been widely used for gene-based therapeutics, but commonly used serotype 5 shows poor transduction efficiency into hematopoietic cells. In this study, we aimed to generate a recombinant adenovirus serotype 5 (rAd5) vector that has a high efficiency in gene transfer to megakaryocytic leukemic cells with anticancer potential. We first modified the rAd5 backbone vector with a chimeric fiber gene of Ad5 and Ad11p (rAd5F11p) to increase the gene delivery efficiency. Then, the nonstructural protein NS1 of human parvovirus B19 (B19V), which induces cell cycle arrest at the G2/M phase and apoptosis, was cloned into the adenoviral shuttle vector. As the expression of parvoviral NS1 protein inhibited Ad replication and production, we engineered the cytomegalovirus (CMV) promoter, which governs NS1 expression, with two tetracycline operator elements (TetO₂). Transfection of the rAd5F11p proviral vectors in Tet repressor-expressing T-REx-293 cells produced rAd in a large quantity. We further evaluated this chimeric rAd5F11p vector in gene delivery in human leukemic cells, UT7/Epo-S1. Strikingly, the novel rAd5F11p-B19NS1-GFP vector, exhibited a transduction efficiency much higher than the original vector, rAd5-B19NS1-GFP, in UT7/Epo-S1 cells, in particular, when they were transduced at a relatively low multiplicity of infection (100 viral genome copies/cell). After the transduction of rAd5F11p-B19NS1-GFP, over 90% of the UT7/Epo-S1 cells were arrested at the G2/M phase, and approximately 40%–50% of the cells were undergoing apoptosis, suggesting the novel rAd5F11P-B19NS1-GFP vector holds a promise in therapeutic potentials of megakaryocytic leukemia. Full article

To mount a strong anti-tumor immune response, non T cell inflamed (cold) tumors may require combination treatment encompassing vaccine strategies preceding checkpoint inhibition. In vivo targeted delivery of tumor-associated antigens (TAA) to dendritic cells (DCs), relying on the natural functions of primary DCs in situ, represents an attractive vaccination strategy. In this study we made use of a full-length MART-1 expressing C/B-chimeric adenoviral vector, consisting of the Ad5 capsid and the Ad3 knob (Ad5/3), which we previously showed to selectively transduce DCs in human skin and lymph nodes. Our data demonstrate that chimeric Ad5/3 vectors encoding TAA, and able to target human DCs in situ, can be used to efficiently induce expansion of functional tumor-specific CD8⁺ effector T cells, either from a naïve T cell pool or from previously primed T cells residing in the melanoma-draining sentinel lymph nodes (SLN). These data support the use of Ad3-knob containing viruses as vaccine vehicles for in vivo delivery. “Off-the-shelf” DC-targeted Ad vaccines encoding TAA could clearly benefit future immunotherapeutic approaches. Full article

A long time has passed since the first clinical trial with adenoviral (Ad) vectors. Despite being very promising, Ad vectors soon revealed their limitations in human clinical trials. The pre-existing immunity, the marked liver tropism and the high toxicity of first generation Ad (FG-Ad) vectors have been the main challenges for the development of new approaches. Significant effort toward the development of genetically and chemically modified adenoviral vectors has enabled researchers to create more sophisticated vectors for gene therapy, with an improved safety profile and a higher transduction ability of different tissues. In this review, we will describe the latest findings in the high-speed, evolving field of genetic and chemical modifications of adenoviral vectors, a field in which different disciplines, such as biomaterial research, virology and immunology, co-operate synergistically to create better gene therapy tools for modern challenges. Full article