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PubMed2026

Recent advances for the pharmaceutical production of highly attenuated poxviruses as viral vector platforms.

INTRODUCTION: Highly attenuated poxviruses serve as potent viral vectors, oncolytic agents, and therapeutic vaccines. They can accommodate and stably maintain a large genomic payload of foreign inserts. Their limited replication in human cells provides an excellent safety profile, but it concomitantly necessitates higher doses of infectious particles for full therapeutic efficacy. AREAS COVERED: We review recent advances in bioprocesses for the pharmaceutical production of poxvirus-based vectors, focusing mainly on the vaccinia virus and the Orf virus. These include upstream processing using highly permissive cell substrates, optimized feeding strategies, and a virus phenotype that facilitates downstream processing. The study explores ongoing challenges and identifies strategies to adapt the downstream process to intensified upstream processes in order to achieve an economic end-to-end production. EXPERT OPINION: For notably increased virus yields of up to 2 log after amplification, we propose to replace classic adsorption chromatography by a collective and continuous purification platform for separating the virus from process-related impurities. Filtration operations facilitate process scalability while reducing volumes, which is beneficial for a flow-through polishing to meet pharmaceutical quality attributes. Combined with artificial intelligence modeling, these advancements alleviate financial pressures on healthcare systems and accelerate the production of novel vaccine candidates for clinical use.

Humans↗

PubMed2026

A randomized trial of viral vector and adjuvanted protein HBV therapeutic vaccine in people with chronic hepatitis B on nucleos(t)ide analogs.

BACKGROUND: This study assessed the safety, efficacy, and immunogenicity of a therapeutic immunization strategy aimed at reaching a functional cure for chronic hepatitis B (CHB), relying on a heterologous prime-boost with viral vectors ChAd155-hIi-HBV and MVA-HBV, combined with sequential or concomitant administration of adjuvanted recombinant HBV proteins (HBc-HBs/AS01B). METHODS: This single-blind, randomized, controlled, first-in-human, phase 1/2 trial enrolled adults aged 18-65 years with HBeAg-negative CHB, virally suppressed on nucleos(t)ide analogs (NAs), with HBsAg >50 IU/mL. Participants received NAs and the following regimens of 4 doses (8-week intervals): sequential administration of ChAd155-hIi-HBV, MVA-HBV, and 2 HBc-HBs/AS01B doses; co-administration of ChAd155-hIi-HBV+HBc-HBs/AS01B, followed by 3 co-administered MVA-HBV+HBc-HBs/AS01B doses; 4 HBc-HBs/AS01B doses; 2 placebo doses followed by ChAd155-hIi-HBV and MVA-HBV administered alone or with HBc-HBs/AS01B; or 4 placebo doses. Safety, efficacy (≥1-log decrease in quantitative (q)HBsAg or HBsAg loss 24 weeks post-dose 4 [day (D)337]), antibody, and T-cell responses were evaluated. RESULTS: In all, 134 participants were vaccinated. Grade 3 solicited adverse events (AEs) (median duration: 2-3 days) were more frequent after co-administration (systemic: 59.3%; administration-site: 33.3%) than sequential administration (systemic: 10.3%; administration-site: 12.8%) of high-dose viral vectors and proteins. No vaccine-related or fatal serious AEs were reported. After 4 doses, no participant had HBsAg loss or ≥1-log decrease in qHBsAg (D337 vs. D1). Co-administration induced the strongest anti-HBs response (73.7% achieved anti-HBs ≥10 mIU/mL 2 weeks post-dose 4 vs. 40.0% after sequential administration). Both sequential and co-administration induced HBc-specific CD4+ and CD8+ T-cell responses, with a prime-boost effect of the viral vectors. CONCLUSIONS: Heterologous prime-boost with ChAd155-hIi-HBV and MVA-HBV, combined with sequential or co-administration of HBc-HBs/AS01B, had an acceptable safety profile, were moderately immunogenic, but no participants showed the expected efficacy outcome.

Humans↗

PubMed2016

Characterization of a novel adeno-associated viral vector with preferential oligodendrocyte tropism.

No adeno-associated virus (AAV) capsid has been described in the literature to exhibit a primary oligodendrocyte tropism when a constitutive promoter drives gene expression, which is a significant barrier for efficient in vivo oligodendrocyte gene transfer. The vast majority of AAV vectors, such as AAV1, 2, 5, 6, 8 or 9, exhibit a dominant neuronal tropism in the central nervous system. However, a novel AAV capsid (Olig001) generated using capsid shuffling and directed evolution was recovered after rat intravenous delivery and subsequent capsid clone rescue, which exhibited a >95% tropism for striatal oligodendrocytes after rat intracranial infusion where a constitutive promoter drove gene expression. Olig001 contains a chimeric mixture of AAV1, 2, 6, 8 and 9, but unlike these parental serotypes after intravenous administration Olig001 has very low affinity for peripheral organs, especially the liver. Furthermore, in mixed glial cell cultures, Olig001 exhibits a 9-fold greater binding when compared with AAV8. This novel oligodendrocyte-preferring AAV vector exhibits characteristics that are a marked departure from previously described AAV serotypes.

Animals↗

PubMed2012

The nontoxic cell cycle modulator indirubin augments transduction of adeno-associated viral vectors and zinc-finger nuclease-mediated gene targeting.

Parameters that regulate or affect the cell cycle or the DNA repair choice between non-homologous end-joining and homology-directed repair (HDR) are excellent targets to enhance therapeutic gene targeting. Here, we have evaluated the impact of five cell-cycle modulating drugs on targeted genome engineering mediated by DNA double-strand break (DSB)-inducing nucleases, such as zinc-finger nucleases (ZFNs). For a side-by-side comparison, we have established four reporter cell lines by integrating a mutated EGFP gene into either three transformed human cell lines or primary umbilical cord-derived mesenchymal stromal cells (UC-MSCs). After treatment with different cytostatic drugs, cells were transduced with adeno-associated virus (AAV) vectors that encode a nuclease or a repair donor to rescue EGFP expression through DSB-induced HDR. We show that transient cell-cycle arrest increased AAV transduction and AAV-mediated HDR up to six-fold in human cell lines and ten-fold in UC-MSCs, respectively. Targeted gene correction was observed in up to 34% of transduced cells. Both the absolute and the relative gene-targeting frequencies were dependent on the cell type, the cytostatic drug, the vector dose, and the nuclease. Treatment of cells with the cyclin-dependent kinase inhibitor indirubin-3'-monoxime was especially promising as this compound combined high stimulatory effects with minimal cytotoxicity. In conclusion, indirubin-3'-monoxime significantly improved AAV transduction and the efficiency of AAV/ZFN-mediated gene targeting and may thus represent a promising compound to enhance DSB-mediated genome engineering in human stem cells, such as UC-MSCs, which hold great promise for future clinical applications.

Blotting, Western↗

PubMed2024

Adeno-Associated Virus Engineering and Load Strategy for Tropism Modification, Immune Evasion and Enhanced Transgene Expression.

Gene therapy aims to add, replace or turn off genes to help treat disease. To date, the US Food and Drug Administration (FDA) has approved 14 gene therapy products. With the increasing interest in gene therapy, feasible gene delivery vectors are necessary for inserting new genes into cells. There are different kinds of gene delivery vectors including viral vectors like lentivirus, adenovirus, retrovirus, adeno-associated virus et al, and non-viral vectors like naked DNA, lipid vectors, polymer nanoparticles, exosomes et al, with viruses being the most commonly used. Among them, the most concerned vector is adeno-associated virus (AAV) because of its safety, natural ability to efficiently deliver gene into cells and sustained transgene expression in multiple tissues. In addition, the AAV genome can be engineered to generate recombinant AAV (rAAV) containing transgene sequences of interest and has been proven to be a safe gene vector. Recently, rAAV vectors have been approved for the treatment of various rare diseases. Despite these approvals, some major limitations of rAAV remain, namely nonspecific tissue targeting and host immune response. Additional problems include neutralizing antibodies that block transgene delivery, a finite transgene packaging capacity, high viral titer used for per dose and high cost. To deal with these challenges, several techniques have been developed. Based on differences in engineering methods, this review proposes three strategies: gene engineering-based capsid modification (capsid modification), capsid surface tethering through chemical conjugation (surface tethering), and other formulations loaded with AAV (virus load). In addition, the major advantages and limitations encountered in rAAV engineering strategies are summarized.

Dependovirus↗

PubMed2026

Virus-induced gene silencing as a tool for functional genomics in weeds: Challenges and future directions.

Virus-induced gene silencing (VIGS) has evolved from a conceptual demonstration of antiviral defense into a pivotal reverse-genetics platform for plant functional genomics. By exploiting engineered DNA- or RNA-based viral vectors, VIGS enables rapid, sequence-specific transcript knockdown through RNA-mediated degradation of target transcripts. Recent refinements in vector design, inoculation strategies, and viral species selection, such as TRV, BSMV, and FoMV, have expanded its application to previously recalcitrant plants, including major crops and emerging weed models. In weeds, functional genomics remains particularly challenging due to high genetic variability, limited genomic resources, and incompatibility with conventional viral vectors and transformation systems. In this context, VIGS provides a tractable approach to investigate genes associated with herbicide resistance, metabolic adaptation, and stress tolerance. Beyond weed biology, its application to studies of immune signaling, hormonal crosstalk, and secondary metabolism highlights VIGS as a versatile biotechnology for elucidating gene function and supporting next-generation strategies in plant improvement and integrated pest management.

Journal Article↗

PubMed2026

Adaptation of lentiviral vectors for viral gene therapy and their impact on host cell biology.

BACKGROUND: Lentiviral vectors (LVVs) are used as a viral gene therapeutic and were derived from human immunodeficiency virus subtype 1 (HIV-1). LVVs are used to deliver and induce the stable expression of transgenes through genome integration. Current clinical LVV delivery systems do not include HIV-1 major accessory genes; however, critical structural and non-structural HIV-1 proteins are encoded by the 4-plasmid combination that composes the 3rd generation LVV transduction systems. LVVs use HIV-1-like mechanisms for viral genome integration and both transgene delivery and expression. LVVs rely on host cell machinery to transcribe and translate transgenes for either knocking down disease-causing genes and/or supplying functional genes in a targeted disease. LVVs integrate into host intronic and intergenic regions due to genomic accessibility, but there are no known biases toward specific target integration motifs. MAIN BODY: Investigation of LVV integration has uncovered the generation of chimeric LVV-host transcripts and altered host transcript splicing patterns. Several Food and Drug Administration (FDA)-approved LVV-derived therapies are used for treating diseases ranging from beta thalassemia to sickle cell anemia. An increasingly popular application of LVV is in the generation of chimeric antigen receptor (CAR) T cell therapies, which change and enhance T cell antigen specificity and effector function in liquid cancers. In November 2023, all CAR T cell therapies were placed under FDA investigation due to higher-than-expected rates of malignant transformation, hospitalization, and death in treated individuals. LVV integrations driving oncogene expression could be a cause for malignancy development. Current methods for resolving LVV integration patterns are technically limited by the sequencing approach applied allowing for only limited characterization of LVV integration profiles and altered host gene regulation. CONCLUSIONS: A comprehensive understanding of LVV integration and its consequences is necessary for understanding how these events influence host cell gene regulation and splicing, possibly identifying tunable variables for enhanced positive clinical outcomes. Here, we review the development of LVV systems, what is known about LVV integration patterns, technologies used to characterize patterns of LVV integration, and what is understood about the subsequent impact on host cell gene regulation and its potential linkage to patient malignancies.

Humans↗

PubMed2026

Transforming Plant Viruses into Vectors for Next-Generation Agriculture-A Review.

Plant viral vectors have evolved from tools for transient gene expression into a versatile platform for precise genetic intervention, offering a rapid, transgene-free alternative to conventional crop transformation. This review critically assesses their engineering for scalable field application, moving beyond foundational techniques like virus-induced gene silencing (VIGS). We highlight how advanced vector design, including deconstructed genomes and synthetic regulatory circuits enhances cargo capacity, specificity, and biosafety. The integration of viral delivery with CRISPR-Cas systems has unlocked virus-induced genome editing (VIGE), base editing, and prime editing, enabling heritable trait modification without tissue culture. However, the transition from proof-of-concept in model plants to robust field technology hinges on overcoming critical bottlenecks: expanding host range through chimeric vectors, ensuring environmental containment, and developing scalable delivery methods such as nano-formulations or adjusted agroinfiltration protocols. We evaluate these delivery routes and emerging synergies with nanobiotechnology for targeted and efficient applications. While challenges in regulation, public perception, and large-scale production persist, the strategic engineering of viral vectors for stability, specificity, and safety positions them as a transformative, next-generation biotechnological input for achieving sustainable crop improvement and protection under changing climatic conditions.

Plant Viruses↗

PubMed2026

Lipid-nanoparticle-mediated base editing of the trabecular meshwork rescues glaucoma in vivo.

Mutations in MYOC, the most common genetic cause of glaucoma, cause misfolded myocilin to accumulate in the endoplasmic reticulum (ER), leading to trabecular meshwork (TM) dysfunction, elevated intraocular pressure, and progressive vision loss. While gene editing offers curative potential, current delivery methods rely on viral vectors, which are limited by inflammation, off-target effects, and poor translatability. Here, we report a nonviral lipid nanoparticle (LNP) platform that enables selective in vivo delivery of mRNA encoding an adenine base editor and single guide RNA (LNP-ABE) to TM cells. A direct comparison of LNP-mCherry with lentiviral GFP revealed that LNPs outperform viral vectors, achieving markedly higher efficiency and greater selectivity for the TM without inducing ocular inflammation. In a Cre-inducible Tg.CreMYOCY437H glaucoma mouse model, LNP-Cre mRNA selectively induced mutant MYOC expression in the TM, faithfully recapitulating key disease features. A single administration of LNP-ABE achieved efficient on-target editing of mutant MYOC, reducing mutant myocilin protein by approximately 46%, decreasing aggregates, alleviating ER stress, and fully rescuing the glaucomatous phenotype in Tg.CreMYOCY437H mice. Importantly, no off-target editing or ocular toxicity was detected. These findings establish LNP-based mRNA delivery as a safe, efficient, and clinically translatable approach for TM-targeted genome editing with broad therapeutic potential in glaucoma.

Animals↗

PubMed2026

Clinical evidence on non-viral CAR-T cell therapies for solid tumors: a scoping review.

BACKGROUND: Chimeric antigen receptor (CAR) T-cell therapy in solid tumors is hindered by the immunosuppressive tumor microenvironment and by toxicities associated with viral-vector manufacturing. Non-viral gene delivery platforms have emerged as a potential alternative, though clinical evidence remains fragmented. METHODS: Following an a priori protocol registered on the Open Science Framework (OSF; https://doi.org/10.17605/OSF.IO/2TPQS) and adhering to JBI/PRISMA-ScR guidelines, a systematic search was conducted across four databases from inception through May 15, 2026. Patient-level data were extracted to describe cellular persistence and clinical outcomes across strictly non-viral delivery platforms. RESULTS: Four early-phase studies met the inclusion criteria, encompassing 28 heavily pretreated patients with metastatic solid tumors. Two non-viral platforms were identified: mRNA electroporation (n=19; intravenous in 13, intratumoral in 6) and the piggyBac transposon system (n=9). Across both mRNA routes, transient CAR-T persistence (<7 days) was observed, with no objective responses (ORR 0%), though disease stabilization yielded a disease control rate (DCR) of 53%; cross-route comparison is limited by differing distribution profiles. The piggyBac system showed longer persistence (~28 days) and a DCR of 78%, including the only documented objective response (ORR 11%). No Grade ≥3 cytokine release syndrome or neurotoxicity was reported in any of the 28 patients, and no tocilizumab or systemic corticosteroids were required. CONCLUSIONS: Within this limited early-phase evidence base, no severe toxicities attributable to non-viral platforms were reported, and the evidence identifies knowledge gaps warranting prospective investigation. mRNA platforms showed transient persistence and disease stabilization in 53% of patients. One partial response was documented with the piggyBac platform in a single patient; however, this outcome cannot be attributed to the delivery platform given simultaneous differences in target antigen, tumor histology, route of administration, and geographic setting. No firm conclusions regarding comparative platform performance can be drawn from this evidence base. SYSTEMATIC REVIEW REGISTRATION: https://doi.org/10.17605/OSF.IO/2TPQS, identifier OSF.IO/2TPQS.

Humans↗

PubMed2026

Research note: Development of a recombinant duck enteritis virus vector expressing DHAV-3 VP1 and DTMUV prM/TE genes.

Duck enteritis virus (DEV) is a promising viral vector for vaccine development. In a previous study, an HDR-CRISPR/Cas9-based strategy was used to generate a recombinant virus, rDEV-DHAV-VP1, by inserting the VP1 gene of duck hepatitis A virus type 3 (DHAV-3) into the UL27/UL26 intergenic region of DEV vaccine strain, resulting in good genetic stability and immunogenicity. In the present study, the same strategy was applied to insert the EGFP gene into the US7/US8 and LORF11/UL55 intergenic regions of a DEV vaccine strain. Among the evaluated insertion sites, the highest level of EGFP expression was observed at the US7/US8 locus, followed by the UL27/UL26 locus. Based on rDEV-DHAV-VP1, the pre-membrane (prM) and truncated envelope (TE) genes of duck Tembusu virus (DTMUV) were further inserted into the US7/US8 locus, resulting in a bivalent recombinant virus, rDEV-VP1-prM/TE. The recombinant virus exhibited growth kinetics comparable to those of the parental virus, while maintaining efficient expression and high genetic stability of the inserted genes. These findings indicate that the HDR-CRISPR/Cas9 system is an efficient strategy for generating stable DEV-based recombinant vectors and provides a promising platform for the development of multivalent vaccines against major duck viral diseases.

CRISPR/Cas9 genome editing↗

PubMed2026

Impact of transfection optimization on adeno-associated virus purification performance and vector quality.

Recombinant adeno-associated virus (rAAV) has shown great promise as a viral vector for gene therapy. However, efficient manufacture of high-quality rAAV to meet clinical demands remains challenging. Here, we optimized rAAV production via a design of experiments (DoE) approach to evaluate the effects of five transfection parameters on vector genome titer, full particle ratio (FE ratio), and viral protein (VP) stoichiometry. The DoE model showed that no single set of transfection conditions could maximize all three responses simultaneously. Subsequently, the materials from DoE-optimized transfections were purified by affinity chromatography followed by anion exchange chromatography (AEX). We observed that AEX recovery declined when capsid loading was high, a situation caused by the combination of high titer with low FE ratio, reducing some of the gains in titer achieved by transfection optimization. Furthermore, AEX had limited capacity to enrich full particles from materials with a very low initial FE ratio. Finally, materials from DoE-optimized transfections showed improved transduction efficiency, which was associated with the ratios of VP1 and VP2 to total VP in the capsid. Overall, this study demonstrates that transfection-derived quality attributes affect purification outcomes and overall vector quality. Our findings highlight the necessity of strategically balancing multiple quality attributes during transfection optimization and provide insights for integrated upstream transfection and downstream purification development.

Adeno-associated virus↗

PubMed2026

Peptide ligands targeting GP64 for the purification of Baculovirus from insect cell lysates.

Autographa californica multiple nucleopolyhedrovirus, known as Baculovirus, is a widely used platform for producing therapeutic proteins and viral vectors. The purity and infectious activity of Baculovirus stocks determine the quality and productivity of recombinant products produced through this system. Current purification strategies suffer from major limitations: centrifugation lacks productivity and scalability; ion-exchange chromatography affords limited selectivity and purity; and the only commercial affinity resin requires harsh elution conditions that significantly reduce functional product recovery. To overcome these limitations, this study introduces the first peptide affinity ligands targeting the baculoviral envelope glycoprotein GP64 for the purification of active Baculovirus particles. We implemented a combinatorial selection workflow based on dual-fluorescence screening of solid-phase peptide libraries to identify 12-mer sequences that bind GP64 and elute Baculovirus under mild conditions (pH 8.5). As the selected ligands are enriched in histidine and tyrosine residues, product release is effected by the combined modulation of pH and ionic strength. Eight candidate peptides (SB1-SB8) were evaluated on Toyopearl and POROS chromatographic resins, demonstrating that matrix chemistry, pore size, and ligand density govern purification performance. The lead peptide SB4 conjugated to POROS resin at ∼10 µmol/mL achieved 81% recovery of infectious virions (transducing units), robust host cell protein reduction (LRV 1.65), and a dynamic binding capacity (DBC10%) of 1.9 × 1010 vg/mL resin. Transmission electron microscopy and multi-angle light scattering confirmed the integrity of purified particles (200 × 50 nm rods with intact nucleocapsids), compared to BacuClear eluates that showed collapsed morphology. The SB4-POROS resin demonstrated storage stability and ∼80% retention of binding capacity over ten purification-regeneration cycles with caustic cleaning. Integration into a three-step downstream process (clarification, affinity capture, and polishing) raised product purity 1,528-fold, from 6.22 × 106 to 9.50 × 109 viral genomes per µg of HCP, while reducing the total HCP burden 1,698-fold, at a cumulative transducing-unit yield of ∼69% relative to the feedstock, establishing SB4-POROS as a promising technology with a favorable projected cost structure for Baculovirus purification.

Affinity chromatography↗

PubMed2026

Therapeutic melanoma vaccines: Platforms, neoantigen strategies, and emerging combination immunotherapies.

Melanoma has emerged as a major focus of cancer immunotherapy research because of its highly immunogenic nature and responsiveness to immune-based treatments. Therapeutic melanoma vaccines are designed to stimulate tumor-specific immune responses through the delivery of Tumor-Associated Antigens (TAAs), Tumor-Specific Antigens (TSAs), and personalized neoantigens. This narrative review provides an overview of current melanoma vaccine strategies, including peptide-based vaccines, dendritic cell vaccines, nucleic acid-based platforms such as mRNA, DNA, and viral vector vaccines. Recent advances in vaccine engineering and tumor genomics have accelerated the development of personalized neoantigen vaccines capable of targeting mutations unique to individual tumors. In parallel, Artificial Intelligence (AI) and Machine Learning (ML) are increasingly being incorporated into neoantigen identification pipelines to improve epitope prediction and optimize vaccine design. Combination strategies involving Immune Checkpoint Inhibitors (ICIs), particularly anti-PD-1 and anti-CTLA-4 therapies, have further enhanced interest in melanoma vaccines by helping overcome tumor-induced immune suppression and augment T-cell activation. In addition to reviewing vaccine mechanisms and emerging technologies, this manuscript examines the evolving clinical trial landscape through analysis of melanoma vaccine studies registered on ClinicalTrials.gov. Although many studies have reported encouraging safety and immunogenicity findings, challenges related to tumor heterogeneity, immune evasion, biomarker selection, and manufacturing complexity continue to limit widespread clinical implementation. Ongoing advances in computational immunology, biomaterial engineering, and precision oncology are expected to further refine melanoma vaccine development and improve therapeutic efficacy. Collectively, these innovations may help establish melanoma vaccines as an increasingly important component of future personalized cancer immunotherapy strategies.

DNA vaccines↗

PubMed2014

Engineering adeno-associated viruses for clinical gene therapy.

Clinical gene therapy has been increasingly successful owing both to an enhanced molecular understanding of human disease and to progressively improving gene delivery technologies. Among these technologies, delivery vectors based on adeno-associated viruses (AAVs) have emerged as safe and effective and, in one recent case, have led to regulatory approval. Although shortcomings in viral vector properties will render extension of such successes to many other human diseases challenging, new approaches to engineer and improve AAV vectors and their genetic cargo are increasingly helping to overcome these barriers.

Capsid↗

PubMed2009

A myocardium tropic adeno-associated virus (AAV) evolved by DNA shuffling and in vivo selection.

To engineer gene vectors that target striated muscles after systemic delivery, we constructed a random library of adeno-associated virus (AAV) by shuffling the capsid genes of AAV serotypes 1 to 9, and screened for muscle-targeting capsids by direct in vivo panning after tail vein injection in mice. After 2 rounds of in vivo selection, a capsid gene named M41 was retrieved mainly based on its high frequency in the muscle and low frequency in the liver. Structural analyses revealed that the AAVM41 capsid is a recombinant of AAV1, 6, 7, and 8 with a mosaic capsid surface and a conserved capsid interior. AAVM41 was then subjected to a side-by-side comparison to AAV9, the most robust AAV for systemic heart and muscle gene delivery; to AAV6, a parental AAV with strong muscle tropism. After i.v. delivery of reporter genes, AAVM41 was found more efficient than AAV6 in the heart and muscle, and was similar to AAV9 in the heart but weaker in the muscle. In fact, the myocardium showed the highest gene expression among all tissues tested in mice and hamsters after systemic AAVM41 delivery. However, gene transfer in non-muscle tissues, mainly the liver, was dramatically reduced. AAVM41 was further tested in a genetic cardiomyopathy hamster model and achieved efficient long-term delta-sarcoglycan gene expression and rescue of cardiac functions. Thus, direct in vivo panning of capsid libraries is a simple tool for the de-targeting and retargeting of viral vector tissue tropisms facilitated by acquisition of desirable sequences and properties.

Animals↗

PubMed2015

Perspective on Adeno-Associated Virus Capsid Modification for Duchenne Muscular Dystrophy Gene Therapy.

Duchenne muscular dystrophy (DMD) is a X-linked, progressive childhood myopathy caused by mutations in the dystrophin gene, one of the largest genes in the genome. It is characterized by skeletal and cardiac muscle degeneration and dysfunction leading to cardiac and/or respiratory failure. Adeno-associated virus (AAV) is a highly promising gene therapy vector. AAV gene therapy has resulted in unprecedented clinical success for treating several inherited diseases. However, AAV gene therapy for DMD remains a significant challenge. Hurdles for AAV-mediated DMD gene therapy include the difficulty to package the full-length dystrophin coding sequence in an AAV vector, the necessity for whole-body gene delivery, the immune response to dystrophin and AAV capsid, and the species-specific barriers to translate from animal models to human patients. Capsid engineering aims at improving viral vector properties by rational design and/or forced evolution. In this review, we discuss how to use the state-of-the-art AAV capsid engineering technologies to overcome hurdles in AAV-based DMD gene therapy.

Animals↗

PubMed2026

Epigenetic regulation of transgenes.

Gene therapy holds significant potential for treating genetic disorders, but the use of viral vectors is limited by factors such as immunogenicity, payload capacity, and high manufacturing costs. Nonviral gene delivery (NVGD) using plasmid DNA presents an attractive alternative; however, it typically provides a limited magnitude or duration of transgene expression. One potential reason for these shortcomings is the host cell's epigenetic regulation mechanisms, which can silence both viral and nonviral transgenes. Specifically, when foreign DNA enters the nucleus, it is detected by nuclear DNA sensors, such as IFI16, which initiate the assembly of a "restrictosome" or nuclear domain 10 (ND10) body. This multiprotein complex contains several components, such as PML, Speckled Proteins (e.g., SP100), DAXX, and ATRX that act as a scaffold for recruiting various epigenetic modifiers that subsequently deposit repressive histone modifications like H3K9me3 and H3K27me3 on the transgene chromatin. These marks induce DNA methylation and the subsequent condensation of plasmids or episomes into heterochromatin, which represses transgene expression. Alternatively, unmethylated CpG motifs in bacterial plasmid DNA can trigger innate immune responses in the cytosol, but this review will specifically focus on the detailed mechanisms of epigenetic regulation responsible for silencing plasmid DNA within the host cell nucleus. Addressing these nuclear defense mechanisms, potentially through strategies that manipulate DNA methylation or inhibit restrictosome activity, is crucial for advancing the development of safe, effective, and long-lasting plasmid viral and non-viral gene therapies.

Epigenesis, Genetic↗