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Novel Viral Vectors for Gene Therapy 2023

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A special issue of Viruses (ISSN 1999-4915). This special issue belongs to the section "Viral Immunology, Vaccines, and Antivirals".

Deadline for manuscript submissions: closed (31 October 2023) | Viewed by 41804

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Dr. Ottmar Herchenröder

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Guest Editor

Institute of Experimental Gene Therapy and Cancer Research, Rostock University Medical Center, 18057 Rostock, Germany
Interests: adenoviruses; lentiviruses; viral vector; cancer research; cellular reprogramming; stem cell research

Prof. Dr. Brigitte Pützer

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Guest Editor

1. Institute of Experimental Gene Therapy and Cancer Research, Rostock University Medical Center, 18057 Rostock, Germany
2. Department Life, Light and Matter, University of Rostock, 18059 Rostock, Germany
Interests: cancer metastasis; immune microenvironment; tumor progression and heterogeneity; drug resistance; adenoviral cell reprogramming technology
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Special Issue Information

Dear Colleagues,

During the ongoing COVID-19 epidemic, virus-derived vectors gained unprecedented popularity as vehicles to vaccinate significant parts of the world’s population against infection with SARS-CoV-2. Facing the challenges posed by nature, along with other novel vaccines, the previously critically viewed viral vector technology has contributed to saving lives. Meanwhile, many more scientists in different areas of medical research have been encouraged to design novel viral vectors for both in vitro and in vivo applications.

This Special Issue of Viruses is dedicated to recent developments in viral vector design and their future, possibly including wider clinical applications, as well as the foreseeable limitations in basic research and patient treatment. Authors who have contributed to the momentum of this field are encouraged to contribute to this Special Issue and both original research and review articles are welcome.

Dr. Ottmar Herchenröder
Prof. Dr. Brigitte Pützer
Guest Editors

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Keywords

viral vector
adenoviral vector
AAV vector
lentiviral vector
inherited disorder
gene therapy
oncolytic virus
vector-based vaccines
targeted gene therapy

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Published Papers (12 papers)

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Editorial

Jump to: Research, Review

2 pages, 136 KiB

Open AccessEditorial

Novel Viral Vectors for Gene Therapy

by Ottmar Herchenröder and Brigitte M. Pützer

Viruses 2024, 16(3), 387; https://doi.org/10.3390/v16030387 - 1 Mar 2024

Viewed by 1607

Abstract

Viral vectors are gene transfer tools assembled from the backbones of naturally occurring viruses [...] Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

Research

Jump to: Editorial, Review

16 pages, 5264 KiB

Open AccessArticle

AAV Vectors Pseudotyped with Capsids from Porcine and Bovine Species Mediate In Vitro and In Vivo Gene Delivery

by Darrick L. Yu, Laura P. van Lieshout, Brenna A. Y. Stevens, Kelsie J. (Jagt) Near, Jenny K. Stodola, Kevin J. Stinson, Durda Slavic and Sarah K. Wootton

Viruses 2024, 16(1), 57; https://doi.org/10.3390/v16010057 - 29 Dec 2023

Cited by 1 | Viewed by 1651

Abstract

Adeno-associated virus (AAV) vectors are among the most widely used delivery vehicles for in vivo gene therapy as they mediate robust and sustained transgene expression with limited toxicity. However, a significant impediment to the broad clinical success of AAV-based therapies is the widespread presence of pre-existing humoral immunity to AAVs in the human population. This immunity arises from the circulation of non-pathogenic endemic human AAV serotypes. One possible solution is to use non-human AAV capsids to pseudotype transgene-containing AAV vector genomes of interest. Due to the low probability of human exposure to animal AAVs, pre-existing immunity to animal-derived AAV capsids should be low. Here, we characterize two novel AAV capsid sequences: one derived from porcine colon tissue and the other from a caprine adenovirus stock. Both AAV capsids proved to be effective transducers of HeLa and HEK293T cells in vitro. In vivo, both capsids were able to transduce the murine nose, lung, and liver after either intranasal or intraperitoneal administration. In addition, we demonstrate that the porcine AAV capsid likely arose from multiple recombination events involving human- and animal-derived AAV sequences. We hypothesize that recurrent recombination events with similar and distantly related AAV sequences represent an effective mechanism for enhancing the fitness of wildtype AAV populations. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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17 pages, 7222 KiB

Open AccessArticle

Insights from the Construction of Adenovirus-Based Vaccine Candidates against SARS-CoV-2: Expecting the Unexpected

by Denice Weklak, Julian Tisborn, Maurin Helen Mangold, Raphael Scheu, Harald Wodrich, Claudia Hagedorn, Franziska Jönsson and Florian Kreppel

Viruses 2023, 15(11), 2155; https://doi.org/10.3390/v15112155 - 25 Oct 2023

Viewed by 1533

Abstract

To contain the spread of the SARS-CoV-2 pandemic, rapid development of vaccines was required in 2020. Rational design, international efforts, and a lot of hard work yielded the market approval of novel SARS-CoV-2 vaccines based on diverse platforms such as mRNA or adenovirus vectors. The great success of these technologies, in fact, contributed significantly to control the pandemic. Consequently, most scientific literature available in the public domain discloses the results of clinical trials and reveals data of efficaciousness. However, a description of processes and rationales that led to specific vaccine design is only partially available, in particular for adenovirus vectors, even though it could prove helpful for future developments. Here, we disclose our insights from the endeavors to design compatible functional adenoviral vector platform expression cassettes for the SARS-CoV-2 spike protein. We observed that contextualizing genes from an ssRNA virus into a DNA virus provides significant challenges. Besides affecting physical titers, expression cassette design of adenoviral vaccine candidates can affect viral propagation and spike protein expression. Splicing of mRNAs was affected, and fusogenicity of the spike protein in ACE2-overexpressing cells was enhanced when the ER retention signal was deleted. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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11 pages, 1083 KiB

Open AccessArticle

A Method to Generate and Rescue Recombinant Adenovirus Devoid of Replication-Competent Particles in Animal-Origin-Free Culture Medium

by Seyyed Mehdy Elahi, Jennifer Jiang, Nazila Nazemi-Moghaddam and Rénald Gilbert

Viruses 2023, 15(11), 2152; https://doi.org/10.3390/v15112152 - 25 Oct 2023

Cited by 1 | Viewed by 1384

Abstract

Adenoviruses are promising vectors for vaccine production and gene therapy. Despite all the efforts in removing animal-derived components such as fetal bovine serum (FBS) during the production of adenovirus vector (AdV), FBS is still frequently employed in the early stages of production. Conventionally, first-generation AdVs (E1 deleted) are generated in different variants of adherent HEK293 cells, and plaque purification (if needed) is performed in adherent cell lines in the presence of FBS. In this study, we generated an AdV stock in SF-BMAdR (A549 cells adapted to suspension culture in serum-free medium). We also developed a limiting dilution method using the same cell line to replace the plaque purification assay. By combining these two technologies, we were able to completely remove the need for FBS from the process of generating and producing AdVs. In addition, we demonstrated that the purified AdV stock is free of any replication-competent adenovirus (RCA). Furthermore, we demonstrated that our limiting dilution method could effectively rescue an AdV from a stock that is highly contaminated with RCA. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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26 pages, 5161 KiB

Open AccessArticle

Modulation of Reoviral Cytolysis (II): Cellular Stemness

by Tarryn Bourhill, Leili Rohani, Mehul Kumar, Pinaki Bose, Derrick Rancourt and Randal N. Johnston

Viruses 2023, 15(7), 1473; https://doi.org/10.3390/v15071473 - 29 Jun 2023

Cited by 4 | Viewed by 1829

Abstract

Oncolytic viruses (OVs) are an emerging cancer therapeutic that are intended to act by selectively targeting and lysing cancerous cells and by stimulating anti-tumour immune responses, while leaving normal cells mainly unaffected. Reovirus is a well-studied OV that is undergoing advanced clinical trials and has received FDA approval in selected circumstances. However, the mechanisms governing reoviral selectivity are not well characterised despite many years of effort, including those in our accompanying paper where we characterize pathways that do not consistently modulate reoviral cytolysis. We have earlier shown that reovirus is capable of infecting and lysing both certain types of cancer cells and also cancer stem cells, and here we demonstrate its ability to also infect and kill healthy pluripotent stem cells (PSCs). This led us to hypothesize that pathways responsible for stemness may constitute a novel route for the modulation of reoviral tropism. We find that reovirus is capable of killing both murine and human embryonic and induced pluripotent stem cells. Differentiation of PSCs alters the cells’ reoviral-permissive state to a resistant one. In a breast cancer cell line that was resistant to reoviral oncolysis, induction of pluripotency programming rendered the cells permissive to cytolysis. Bioinformatic analysis indicates that expression of the Yamanaka pluripotency factors may be associated with regulating reoviral selectivity. Mechanistic insights from these studies will be useful for the advancement of reoviral oncolytic therapy. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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15 pages, 6994 KiB

Open AccessArticle

Modulation of Reoviral Cytolysis (I): Combination Therapeutics

by Yoshinori Mori, Sandra G. Nishikawa, Andreea R. Fratiloiu, Mio Tsutsui, Hiromi Kataoka, Takashi Joh and Randal N. Johnston

Viruses 2023, 15(7), 1472; https://doi.org/10.3390/v15071472 - 29 Jun 2023

Cited by 1 | Viewed by 1571

Abstract

Patients with stage IV gastric cancer suffer from dismal outcomes, a challenge especially in many Asian populations and for which new therapeutic options are needed. To explore this issue, we used oncolytic reovirus in combination with currently used chemotherapeutic drugs (irinotecan, paclitaxel, and docetaxel) for the treatment of gastric and other gastrointestinal cancer cells in vitro and in a mouse model. Cell viability in vitro was quantified by WST-1 assays in human cancer cell lines treated with reovirus and/or chemotherapeutic agents. The expression of reovirus protein and caspase activity was determined by flow cytometry. For in vivo studies, athymic mice received intratumoral injections of reovirus in combination with irinotecan or paclitaxel, after which tumor size was monitored. In contrast to expectations, we found that reoviral oncolysis was only poorly correlated with Ras pathway activation. Even so, the combination of reovirus with chemotherapeutic agents showed synergistic cytopathic effects in vitro, plus enhanced reovirus replication and apoptosis. In vivo experiments showed that reovirus alone can reduce tumor size and that the combination of reovirus with chemotherapeutic agents enhances this effect. Thus, we find that oncolytic reovirus therapy is effective against gastric cancer. Moreover, the combination of reovirus and chemotherapeutic agents synergistically enhanced cytotoxicity in human gastric cancer cell lines in vitro and in vivo. Our data support the use of reovirus in combination with chemotherapy in further clinical trials, and highlight the need for better biomarkers for reoviral oncolytic responsiveness. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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11 pages, 1441 KiB

Open AccessArticle

Adeno-Associated Virus-like Particles’ Response to pH Changes as Revealed by nES-DMA

by Samuele Zoratto, Thomas Heuser, Gernot Friedbacher, Robert Pletzenauer, Michael Graninger, Martina Marchetti-Deschmann and Victor U. Weiss

Viruses 2023, 15(6), 1361; https://doi.org/10.3390/v15061361 - 13 Jun 2023

Cited by 2 | Viewed by 1958

Abstract

Gas-phase electrophoresis on a nano-Electrospray Gas-phase Electrophoretic Mobility Molecular Analyzer (nES GEMMA) separates single-charged, native analytes according to the surface-dry particle size. A volatile electrolyte, often ammonium acetate, is a prerequisite for electrospraying. Over the years, nES GEMMA has demonstrated its unique capability to investigate (bio-)nanoparticle containing samples in respect to composition, analyte size, size distribution, and particle numbers. Virus-like particles (VLPs), being non-infectious vectors, are often employed for gene therapy applications. Focusing on adeno-associated virus 8 (AAV8) based VLPs, we investigated the response of these bionanoparticles to pH changes via nES GEMMA as ammonium acetate is known to exhibit these changes upon electrospraying. Indeed, slight yet significant differences in VLP diameters in relation to pH changes are found between empty and DNA-cargo-filled assemblies. Additionally, filled VLPs exhibit aggregation in dependence on the applied electrolyte’s pH, as corroborated by atomic force microscopy. In contrast, cryogenic transmission electron microscopy did not relate to changes in the overall particle size but in the substantial particle’s shape based on cargo conditions. Overall, we conclude that for VLP characterization, the pH of the applied electrolyte solution has to be closely monitored, as variations in pH might account for drastic changes in particles and VLP behavior. Likewise, extrapolation of VLP behavior from empty to filled particles has to be carried out with caution. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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12 pages, 9599 KiB

Open AccessArticle

Engineering Adenoviral Vectors with Improved GBM Selectivity

by Emily A. Bates, Charlotte Lovatt, Alice R. Plein, James A. Davies, Florian A. Siebzehnrubl and Alan L. Parker

Viruses 2023, 15(5), 1086; https://doi.org/10.3390/v15051086 - 28 Apr 2023

Cited by 8 | Viewed by 2461

Abstract

Glioblastoma (GBM) is the most common and aggressive adult brain cancer with an average survival rate of around 15 months in patients receiving standard treatment. Oncolytic adenovirus expressing therapeutic transgenes represent a promising alternative treatment for GBM. Of the many human adenoviral serotypes described to date, adenovirus 5 (HAdV-C5) has been the most utilised clinically and experimentally. However, the use of Ad5 as an anti-cancer agent may be hampered by naturally high seroprevalence rates to HAdV-C5 coupled with the infection of healthy cells via native receptors. To explore whether alternative natural adenoviral tropisms are better suited to GBM therapeutics, we pseudotyped an HAdV-C5-based platform using the fibre knob protein from alternative serotypes. We demonstrate that the adenoviral entry receptor coxsackie, adenovirus receptor (CAR) and CD46 are highly expressed by both GBM and healthy brain tissue, whereas Desmoglein 2 (DSG2) is expressed at a low level in GBM. We demonstrate that adenoviral pseudotypes, engaging CAR, CD46 and DSG2, effectively transduce GBM cells. However, the presence of these receptors on non-transformed cells presents the possibility of off-target effects and therapeutic transgene expression in healthy cells. To enhance the specificity of transgene expression to GBM, we assessed the potential for tumour-specific promoters hTERT and survivin to drive reporter gene expression selectively in GBM cell lines. We demonstrate tight GBM-specific transgene expression using these constructs, indicating that the combination of pseudotyping and tumour-specific promoter approaches may enable the development of efficacious therapies better suited to GBM. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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22 pages, 2084 KiB

Open AccessArticle

The Optimized γ-Globin Lentiviral Vector GGHI-mB-3D Leads to Nearly Therapeutic HbF Levels In Vitro in CD34⁺ Cells from Sickle Cell Disease Patients

by Ekati Drakopoulou, Maria Georgomanoli, Carsten W. Lederer, Fottes Panetsos, Marina Kleanthous, Ersi Voskaridou, Dimitrios Valakos, Eleni Papanikolaou and Nicholas P. Anagnou

Viruses 2022, 14(12), 2716; https://doi.org/10.3390/v14122716 - 5 Dec 2022

Cited by 3 | Viewed by 2165

Abstract

We have previously demonstrated that both the original γ-globin lentiviral vector (LV) GGHI and the optimized GGHI-mB-3D LV, carrying the novel regulatory elements of the 3D HPFH-1 enhancer and the 3’ β-globin UTR, can significantly increase HbF production in thalassemic CD34⁺ cells and ameliorate the disease phenotype in vitro. In the present study, we investigated whether the GGHI-mB-3D vector can also exhibit an equally therapeutic effect, following the transduction of sickle cell disease (SCD) CD34⁺ cells at MOI 100, leading to HbF increase coupled with HbS decrease, and thus, to phenotype improvement in vitro. We show that GGHI-mB-3D LV can lead to high and potentially therapeutic HbF levels, reaching a mean 2-fold increase to a mean value of VCN/cell of 1.0 and a mean transduction efficiency of 55%. Furthermore, this increase was accompanied by a significant 1.6-fold HbS decrease, a beneficial therapeutic feature for SCD. In summary, our data demonstrate the efficacy of the optimized γ-globin lentiviral vector to improve the SCD phenotype in vitro, and highlights its potential use in future clinical SCD trials. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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Review

Jump to: Editorial, Research

13 pages, 273 KiB

Open AccessReview

Suitable Disinfectants with Proven Efficacy for Genetically Modified Viruses and Viral Vectors

by Maren Eggers, Ingeborg Schwebke, Johannes Blümel, Franziska Brandt, Helmut Fickenscher, Jürgen Gebel, Nils Hübner, Janis A. Müller, Holger F. Rabenau, Ingrid Rapp, Sven Reiche, Eike Steinmann, Jochen Steinmann, Paula Zwicker and Miranda Suchomel

Viruses 2023, 15(11), 2179; https://doi.org/10.3390/v15112179 - 30 Oct 2023

Cited by 2 | Viewed by 2111

Abstract

Viral disinfection is important for medical facilities, the food industry, and the veterinary field, especially in terms of controlling virus outbreaks. Therefore, standardized methods and activity levels are available for these areas. Usually, disinfectants used in these areas are characterized by their activity against test organisms (i.e., viruses, bacteria, and/or yeasts). This activity is usually determined using a suspension test in which the test organism is incubated with the respective disinfectant in solution to assess its bactericidal, yeasticidal, or virucidal activity. In addition, carrier methods that more closely reflect real-world applications have been developed, in which microorganisms are applied to the surface of a carrier (e.g., stainless steel frosted glass, or polyvinyl chloride (PVC)) and then dried. However, to date, no standardized methods have become available for addressing genetically modified vectors or disinfection-resistant oncolytic viruses such as the H1-parvovirus. Particularly, such non-enveloped viruses, which are highly resistant to disinfectants, are not taken into account in European standards. This article proposes a new activity claim known as “virucidal activity PLUS”, summarizes the available methods for evaluating the virucidal activity of chemical disinfectants against genetically modified organisms (GMOs) using current European standards, including the activity against highly resistant parvoviridae such as the adeno-associated virus (AAV), and provides guidance on the selection of disinfectants for pharmaceutical manufacturers, laboratories, and clinical users. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

25 pages, 5790 KiB

Open AccessReview

Innate Immune Response to Viral Vectors in Gene Therapy

by Yixuan Wang and Wenwei Shao

Viruses 2023, 15(9), 1801; https://doi.org/10.3390/v15091801 - 24 Aug 2023

Cited by 8 | Viewed by 4248

Abstract

Viral vectors play a pivotal role in the field of gene therapy, with several related drugs having already gained clinical approval from the EMA and FDA. However, numerous viral gene therapy vectors are currently undergoing pre-clinical research or participating in clinical trials. Despite advancements, the innate response remains a significant barrier impeding the clinical development of viral gene therapy. The innate immune response to viral gene therapy vectors and transgenes is still an important reason hindering its clinical development. Extensive studies have demonstrated that different DNA and RNA sensors can detect adenoviruses, adeno-associated viruses, and lentiviruses, thereby activating various innate immune pathways such as Toll-like receptor (TLR), cyclic GMP-AMP synthase–stimulator of interferon genes (cGAS-STING), and retinoic acid-inducible gene I–mitochondrial antiviral signaling protein (RLR-MAVS). This review focuses on elucidating the mechanisms underlying the innate immune response induced by three widely utilized viral vectors: adenovirus, adeno-associated virus, and lentivirus, as well as the strategies employed to circumvent innate immunity. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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35 pages, 951 KiB

Open AccessReview

Viral Vectors in Gene Therapy: Where Do We Stand in 2023?

by Kenneth Lundstrom

Viruses 2023, 15(3), 698; https://doi.org/10.3390/v15030698 - 7 Mar 2023

Cited by 58 | Viewed by 17107

Abstract

Viral vectors have been used for a broad spectrum of gene therapy for both acute and chronic diseases. In the context of cancer gene therapy, viral vectors expressing anti-tumor, toxic, suicide and immunostimulatory genes, such as cytokines and chemokines, have been applied. Oncolytic viruses, which specifically replicate in and kill tumor cells, have provided tumor eradication, and even cure of cancers in animal models. In a broader meaning, vaccine development against infectious diseases and various cancers has been considered as a type of gene therapy. Especially in the case of COVID-19 vaccines, adenovirus-based vaccines such as ChAdOx1 nCoV-19 and Ad26.COV2.S have demonstrated excellent safety and vaccine efficacy in clinical trials, leading to Emergency Use Authorization in many countries. Viral vectors have shown great promise in the treatment of chronic diseases such as severe combined immunodeficiency (SCID), muscular dystrophy, hemophilia, β-thalassemia, and sickle cell disease (SCD). Proof-of-concept has been established in preclinical studies in various animal models. Clinical gene therapy trials have confirmed good safety, tolerability, and therapeutic efficacy. Viral-based drugs have been approved for cancer, hematological, metabolic, neurological, and ophthalmological diseases as well as for vaccines. For example, the adenovirus-based drug Gendicine^® for non-small-cell lung cancer, the reovirus-based drug Reolysin^® for ovarian cancer, the oncolytic HSV T-VEC for melanoma, lentivirus-based treatment of ADA-SCID disease, and the rhabdovirus-based vaccine Ervebo against Ebola virus disease have been approved for human use. Full article

(This article belongs to the Special Issue Novel Viral Vectors for Gene Therapy 2023)

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