Optimization in the expression of ASFV proteins for the development of subunit vaccines using poxviruses as delivery vectors

African swine fever virus (ASFV) causes a highly contagious hemorrhagic disease that affects domestic pig and Eurasian wild boar populations. To date, no safe and efficacious treatment or vaccine against ASF is available. Nevertheless, there are several reports of protection elicited by experimental vaccines based on live attenuated ASFV and some levels of protection and reduced viremia in other approaches such as DNA, adenovirus, baculovirus, and vaccinia-based vaccines. Current ASF subunit vaccine research focuses mainly on delivering protective antigens and antigen discovery within the ASFV genome. However, due to the complex nature of ASFV, expression vectors need to be optimized to improve their immunogenicity. Therefore, in the present study, we constructed several recombinant MVA vectors to evaluate the efficiency of different promoters and secretory signal sequences in the expression and immunogenicity of the p30 protein from ASFV. Overall, the natural poxvirus PrMVA13.5L promoter induced high levels of both p30 mRNA and specific anti-p30 antibodies in mice. In contrast, the synthetic PrS5E promoter and the S E/L promoter linked to a secretory signal showed lower mRNA levels and antibodies. These findings indicate that promoter selection may be as crucial as the antigen used to develop ASFV subunit vaccines using MVA as the delivery vector.

Optimization in the Expression of ASFV Proteins for the Development of Subunit Vaccines Using Poxviruses As Delivery Vectors.

African swine fever virus (ASFV) causes a highly contagious hemorrhagic disease that affects domestic pig and Eurasian wild boar populations. To date, no safe and efficacious treatment or vaccine against ASF is available. Nevertheless, there are several reports of protection elicited by experimental vaccines based on live attenuated ASFV and some levels of protection and reduced viremia in other approaches such as DNA, adenovirus, baculovirus, and vaccinia-based vaccines. Current ASF subunit vaccine research focuses mainly on delivering protective antigens and antigen discovery within the ASFV genome. However, due to the complex nature of ASFV, expression vectors need to be optimized to improve their immunogenicity. Therefore, in the present study, we constructed several recombinant MVA vectors to evaluate the efficiency of different promoters and secretory signal sequences in the expression and immunogenicity of the p30 protein from ASFV. Overall, the natural poxvirus PrMVA13.5L promoter induced high levels of both p30 mRNA and specific anti-p30 antibodies in mice. In contrast, the synthetic PrS5E promoter and the S E/L promoter linked to a secretory signal showed lower mRNA levels and antibodies. These findings indicate that promoter selection may be as crucial as the antigen used to develop ASFV subunit vaccines using MVA as the delivery vector.

Development and Characterization of High Efficacy Cell-Penetrating Peptide via Modulation of the Histidine and Arginine Ratio for Gene Therapy

Cell-penetrating peptides (CPPs), as non-viral gene delivery vectors, are considered with lower immunogenic response, and safer and higher gene capacity than viral systems. In our previous study, a CPP peptide called RALA (arginine rich) presented desirable transfection efficacy and owns a potential clinic use. It is believed that histidine could enhance the endosome escaping ability of CPPs, yet RALA peptide contains only one histidine in each chain. In order to develop novel superior CPPs, by using RALA as a model, we designed a series of peptides named HALA (increased histidine ratio). Both plasmid DNA (pDNA) and siRNA transfection results on three cell lines revealed that the transfection efficacy is better when histidine replacements were on the C-terminal instead of on the N-terminal, and two histidine replacements are superior to three. By investigating the mechanism of endocytosis of the pDNA nanocomplexes, we discovered that there were multiple pathways that led to the process and caveolae played the main role. During the screening, we discovered a novel peptide-HALA2 of high cellular transfection efficacy, which may act as an exciting gene delivery vector for gene therapy. Our findings also bring new insights on the development of novel robust CPPs.

Synthesis and characterization of vitamin D3-functionalized carbon dots for CRISPR/Cas9 delivery

Aim: To develop a novel nanovector for the delivery of genetic fragments and CRISPR/Cas9 systems in particular. Materials & methods: Vitamin D3-functionalized carbon dots (D/CDs) fabricated using one-step microwave-aided methods were characterized by different microscopic and spectroscopic techniques. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide assay and flow cytometry were employed to determine the cell viability and transfection efficiency. Results: D/CDs transfected CRISPR plasmid in various cell lines with high efficiency while maintaining their remarkable efficacy at high serum concentration and low plasmid doses. They also showed great potential for the green fluorescent protein disruption by delivering two different types of CRISPR/Cas9 systems. Conclusion: Given their high efficiency and safety, D/CDs provide a versatile gene-delivery vector for clinical applications.

Advances in Oral Chitosan Based Nano Delivery System for colon Targeted Drug Delivery in Inflammatory Bowel Disease

Nanomaterials can be used as drug carriers with multiple features, including target delivery triggered by environmental, pH, thermal responses, enhanced biocompatibility, and the ability to cross the blood-brain barrier. Chitosan (CS) is a natural polysaccharide largely obtained from marine crustaceans. It provides drug delivery vector for therapeutic CS and diagnostic CS, owing to its biocompatibility, biodegradability, low toxicity, and structural variability. Derivatives of CS such as quaternized CS, thiolated CS and carboxylated CS have enhanced its effectiveness in oral absorption of macromolecular drugs. This review discusses different forms of nanomaterials generated from CS and its derivatives for controlled drug delivery.

Dendrimer-Modified Gold Nanorods As A Platform For Combinational Gene Therapy And Photothermal Therapy of Tumors

Background: The exploitation of novel nanomaterials combining diagnostic and therapeutic functionalities within one single nanoplatform is challenging for tumor theranostics. Methods: We synthesized dendrimer-modified gold nanorods for combinational gene therapy and photothermal therapy (PTT) of colon cancer. Poly(amidoamine) dendrimers (PAMAM, G3) grafted gold nanorods were modified with GX1 peptide (a cyclic 7-mer peptide, CGNSNPKSC). The obtained Au NR@PAMAM-GX1 are performed as a gene delivery vector to gene (FAM172A, regulates the proliferation and apoptosis of colon cancer cells) for the combination of photothermal therapy (PTT) and gene therapy of Colon cancer cells (HCT-8 cells). In addition, the CT imaging function of Au NR can provide imaging evidence for the diagnosis of colon cancer.Results: The results display that Au NR@PAMAM-GX1 can specifically deliver FAM172A to cancer cells with excellent transfection efficiency. The HCT-8 cells treated with the Au NR@PAMAM-GX1/FAM172A under laser irradiation have a viability of 20.45%, which is much lower than the survival rate of other single-mode PTT treatment or single-mode gene therapy. Furthermore, animal experiment results confirm that Au NR@PAMAM-GX1/FAM172A complexes can achieve tumor thermal imaging, PTT and gene therapy after tail vein injection. Conclusion: Our findings demonstrate that the synthesized Au NR@PAMAM-GX1 offer a facile platform to exert antitumor and improve the diagnostic level of tumor.

Nerve growth factor loaded macrophage-derived nanovesicles for inhibiting neuronal apoptosis after spinal cord injury

Spinal cord injury (SCI) is an extremely destructive central nervous system lesion. Studies have shown that NGF can promote nerve regeneration after SCI. However, it cannot produce the desired effect due to its stability in the body and is difficulty in passing through the blood-brain barrier. In this study, we prepared nanovesicles derived from macrophage membrane encapsulating NGF (NGF-NVs) as a drug carrier for the treatment of SCI. Cell experiments showed that NGF-NVs were effectively taken up by PC12 cells and inhibited neuronal apoptosis. In vivo imaging experiments, a large quantity of NGF was delivered to the injured site with the aid of the good targeting of NVs. In animal experiments, NGF-NVs improved the survival of neurons by significantly activating the PI3K/AKT signaling pathway and had good behavioral and histological recovery effects after SCI. Therefore, NVs are a potential drug delivery vector for SCI therapy.

Nanoformulations can be utilized to deliver effective siRNA to tumor cells to decrease gene expression

siRNA-based gene therapy has gained interest in recent decades. Nanocarriers can be used to deliver rapid and efficient siRNA to tumor cells, suppressing gene expression. Some RNAi-based therapies are presently undergoing clinical trials. Clinical trials to preserve and distribute siRNA are increasingly focused on nanoformulations, and their safety has been promising. How nanocarriers avoid immune clearance and extend their circulation in the body is a key study topic for scientists.Most research relies on EPR-mediated passive targeting. However, as the MPS system sequesters most nanocarriers, the E PR effect can not be administered successfully in mice, which is a big human concern. Novel targeted approaches are desperately needed to accumulate siRNA in tumor tissues. In vitro and animal models, a number of TME-responsive siRNA nanocarriers were examined. Clinical nanomedicine is generally simple in structure and easy to mass-produce while preserving safety and effectiveness.It will require the collaborative efforts of materials scientists, basic medical scientists, and clinicians to develop the design of the ideal siRNA delivery vector, the authors suggest. The authors suggest that multimodal combination therapy has a stronger therapeutic impact on malignancies than a single therapy paradigm and might be "over-designed" for therapeutic usage. The study was published in Cancer Research.

Understanding why the same gene delivery vector behaves differently in different cell types is essential for developing more adaptable transfection systems

Since their origin, non-viral gene delivery reagents have evolved into a variety of effective delivery reagents with a variety of components and designs, and are widely used in gene therapy and gene engineering. A flood of successful commercial gene delivery reagents has also developed, and PEI has emerged as the "gold standard" for the industry. On the other hand, their transfection efficiency must be enhanced and their cell toxicity must be reduced. In recent years, toxicity, efficiency and targeted investigations have progressed. In addition to creating and manufacturing reagents with reduced toxicity and higher efficiency, polypeptides that stimulate cell membrane perforation and tiny molecular compounds that can better compress pDNA, as well as various combinations with liposomes or polymer vectors, have demonstrated improved outcomes. However, most of these freshly created delivery vector reagents are still under investigation, and others require additional refinement to achieve high transfection efficiency and minimum toxicity. The processes behind the effects of various gene delivery reagents, genes, and drugs entering cells, as well as their transit, escape, and cell metabolism, are also unclear. This requires improving relevant research. Understanding why the same reagent reacts differently to different cell types is crucial to creating more adaptive transfection reagents for different cell lines. This is suggested because different cells have different growth cycles. Because of their weak proliferation capacity, primordial cells, for example, are harder to replicate.Artificial intelligence, real-world and virtual-world integration technology, big data, multiomics technology, and signal pathway research have all achieved substantial breakthroughs in recent years, and novel transfection reagents and drug delivery technologies are predicted to continue. It is worth examining how to take advantage of the scientific and high-efficiency benefits that new technology provides for research and how to solve the issues given by the in-depth examination of the selection and mechanism of action of novel composite materials in vector reagent creation.

Charge-alteration-based approaches can address the evolving needs of nucleic acid-based gene therapy, charge reversal techniques are also promising

Recent approvals of RNAi-based medications have rekindled interest in the industry, possibly strengthening the power of the therapeutic platform. Many groundbreaking research endeavors have offered new carrier architectures for carrying nucleic acid payloads. Because nucleic acids are charged negatively, utilizing complementary positively charged compounds sounds like a smart approach. The presence of such a cationic charge in the delivery vector, however, is a serious problem, increasing cytotoxicity. Charge-alteration-based approaches can address the evolving needs of nucleic acid-based therapies. Medication delivery platforms have become significantly smarter and safer.The research focuses on exciting recent advancements in management-switching technology. This study summarizes some of these fascinating possibilities, written by authors with special attention to the underlying physics of charge conversion processes. Charge reversal techniques are fairly new and are still in the early development phases. Translating such ideas would require significant investigation, including studies of safety, efficacy, and immunogenicity using relevant disease models.