scholarly journals Riboswitches for Controlled Expression of Therapeutic Transgenes Delivered by Adeno-Associated Viral Vectors

2021 ◽  
Vol 14 (6) ◽  
pp. 554
Author(s):  
Zachary J. Tickner ◽  
Michael Farzan
Keyword(s):  
Gene Therapy ◽  
Animal Models ◽  
Mammalian Cells ◽  
Viral Vectors ◽  
Adeno Associated Virus ◽  
Gene Therapies ◽  
Expression Control ◽  
Transgene Delivery ◽  
Improved Performance

Vectors developed from adeno-associated virus (AAV) are powerful tools for in vivo transgene delivery in both humans and animal models, and several AAV-delivered gene therapies are currently approved for clinical use. However, AAV-mediated gene therapy still faces several challenges, including limited vector packaging capacity and the need for a safe, effective method for controlling transgene expression during and after delivery. Riboswitches, RNA elements which control gene expression in response to ligand binding, are attractive candidates for regulating expression of AAV-delivered transgene therapeutics because of their small genomic footprints and non-immunogenicity compared to protein-based expression control systems. In addition, the ligand-sensing aptamer domains of many riboswitches can be exchanged in a modular fashion to allow regulation by a variety of small molecules, proteins, and oligonucleotides. Riboswitches have been used to regulate AAV-delivered transgene therapeutics in animal models, and recently developed screening and selection methods allow rapid isolation of riboswitches with novel ligands and improved performance in mammalian cells. This review discusses the advantages of riboswitches in the context of AAV-delivered gene therapy, the subsets of riboswitch mechanisms which have been shown to function in human cells and animal models, recent progress in riboswitch isolation and optimization, and several examples of AAV-delivered therapeutic systems which might be improved by riboswitch regulation.

Gene therapy comes of age

Science ◽  
2018 ◽  
Vol 359 (6372) ◽  
pp. eaan4672 ◽  
Author(s):  
Cynthia E. Dunbar ◽  
Katherine A. High ◽  
J. Keith Joung ◽  
Donald B. Kohn ◽  
Keiya Ozawa ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Viral Vectors ◽  
The United States ◽  
Genetically Engineered ◽  
Drug Status ◽  
Hematopoietic Stem ◽  
Gene Therapies ◽  
Engineered T Cells ◽  
Near Future

After almost 30 years of promise tempered by setbacks, gene therapies are rapidly becoming a critical component of the therapeutic armamentarium for a variety of inherited and acquired human diseases. Gene therapies for inherited immune disorders, hemophilia, eye and neurodegenerative disorders, and lymphoid cancers recently progressed to approved drug status in the United States and Europe, or are anticipated to receive approval in the near future. In this Review, we discuss milestones in the development of gene therapies, focusing on direct in vivo administration of viral vectors and adoptive transfer of genetically engineered T cells or hematopoietic stem cells. We also discuss emerging genome editing technologies that should further advance the scope and efficacy of gene therapy approaches.

scholarly journals Weighing the DNA content of Adeno-Associated Virus vectors with zeptogram precision using nanomechanical resonators

2021 ◽  
Author(s):  
Georgios Katsikis ◽  
Iris E Hwang ◽  
Wade Wang ◽  
Vikas S Bhat ◽  
Nicole L McIntosh ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Quality Control ◽  
Viral Vectors ◽  
Analytical Ultracentrifugation ◽  
Vaccine Development ◽  
Turnaround Time ◽  
Charge Detection ◽  
Adeno Associated Virus ◽  
Nanomechanical Resonators

Quantifying the composition of viral vectors used in vaccine development and gene therapy is critical for assessing their functionality. Adeno-Associated Virus (AAV) vectors, which are the most widely used viral vectors for in-vivo gene therapy, are typically characterized using PCR, ELISA, and Analytical Ultracentrifugation which require laborious protocols or hours of turnaround time. Emerging methods such as Charge-Detection Mass Spectroscopy, Static Light Scattering, and Mass Photometry offer turnaround times of minutes for measuring AAV mass, but mostly require purified AAV-based reference materials for calibration. Here, we demonstrate a method for using Suspended Nanomechanical Resonators (SNR) to directly measure both AAV mass and aggregation from a few microliters of sample within minutes. We achieve a resolution near 10 zeptograms which corresponds to 1% of the genome holding capacity of the AAV capsid. Our results show the potential of our method for providing real-time quality control of viral vectors during biomanufacturing.

scholarly journals The Structure of an AAV5-AAVR Complex at 2.5 Å Resolution: Implications for Cellular Entry and Immune Neutralization of AAV Gene Therapy Vectors

Viruses ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 1326
Author(s):  
Mark A. Silveria ◽  
Edward E. Large ◽  
Grant M. Zane ◽  
Tommi A. White ◽  
Michael S. Chapman
Keyword(s):  
Gene Therapy ◽  
Binding Sites ◽  
Polycystic Kidney ◽  
Adeno Associated Virus ◽  
Gene Therapies ◽  
The Us ◽  
Gene Therapy Vectors ◽  
Cellular Entry ◽  
Resolution Structure

Adeno-Associated Virus is the leading vector for gene therapy. Although it is the vector for all in vivo gene therapies approved for clinical use by the US Food and Drug Administration, its biology is still not yet fully understood. It has been shown that different serotypes of AAV bind to their cellular receptor, AAVR, in different ways. Previously we have reported a 2.4Å structure of AAV2 bound to AAVR that shows ordered structure for only one of the two AAVR domains with which AAV2 interacts. In this study we present a 2.5Å resolution structure of AAV5 bound to AAVR. AAV5 binds to the first polycystic kidney disease (PKD) domain of AAVR that was not ordered in the AAV2 structure. Interactions of AAV5 with AAVR are analyzed in detail, and the implications for AAV2 binding are explored through molecular modeling. Moreover, we find that binding sites for the antibodies ADK5a, ADK5b, and 3C5 on AAV5 overlap with the binding site of AAVR. These insights provide a structural foundation for development of gene therapy agents to better evade immune neutralization without disrupting cellular entry.

Non-Viral Vectors for Gene Delivery

2018 ◽  
Vol 9 (1) ◽  
pp. 4-11 ◽  
Author(s):  
Aparna Bansal ◽  
Himanshu
Keyword(s):  
Gene Therapy ◽  
Viral Vectors ◽  
Transfection Efficiency ◽  
Gene Transfection ◽  
Expression System ◽  
Target Cells ◽  
Specific Expression ◽  
Naked Dna

Introduction: Gene therapy has emerged out as a promising therapeutic pave for the treatment of genetic and acquired diseases. Gene transfection into target cells using naked DNA is a simple and safe approach which has been further improved by combining vectors or gene carriers. Both viral and non-viral approaches have achieved a milestone to establish this technique, but non-viral approaches have attained a significant attention because of their favourable properties like less immunotoxicity and biosafety, easy to produce with versatile surface modifications, etc. Literature is rich in evidences which revealed that undoubtedly, non–viral vectors have acquired a unique place in gene therapy but still there are number of challenges which are to be overcome to increase their effectiveness and prove them ideal gene vectors. Conclusion: To date, tissue specific expression, long lasting gene expression system, enhanced gene transfection efficiency has been achieved with improvement in delivery methods using non-viral vectors. This review mainly summarizes the various physical and chemical methods for gene transfer in vitro and in vivo.

scholarly journals Planet of the AAVs: The Spinal Cord Injury Episode

Biomedicines ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 613
Author(s):  
Katerina Stepankova ◽  
Pavla Jendelova ◽  
Lucia Machova Urdzikova
Keyword(s):  
Spinal Cord ◽  
Gene Therapy ◽  
Spinal Cord Injury ◽  
Cell Types ◽  
Specific Cell ◽  
Adeno Associated Virus ◽  
Adequate Treatment ◽  
Cord Injury ◽  
The Central Nervous System ◽  
Transgene Delivery

The spinal cord injury (SCI) is a medical and life-disrupting condition with devastating consequences for the physical, social, and professional welfare of patients, and there is no adequate treatment for it. At the same time, gene therapy has been studied as a promising approach for the treatment of neurological and neurodegenerative disorders by delivering remedial genes to the central nervous system (CNS), of which the spinal cord is a part. For gene therapy, multiple vectors have been introduced, including integrating lentiviral vectors and non-integrating adeno-associated virus (AAV) vectors. AAV vectors are a promising system for transgene delivery into the CNS due to their safety profile as well as long-term gene expression. Gene therapy mediated by AAV vectors shows potential for treating SCI by delivering certain genetic information to specific cell types. This review has focused on a potential treatment of SCI by gene therapy using AAV vectors.

scholarly journals Pseudotyping Lentiviral Vectors: When the Clothes Make the Virus

Viruses ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 1311 ◽  
Author(s):  
Alexis Duvergé ◽  
Matteo Negroni
Keyword(s):  
Gene Therapy ◽  
Genetic Modification ◽  
Viral Vectors ◽  
Molecular Level ◽  
Lentiviral Vectors ◽  
Human Cells ◽  
Surface Proteins ◽  
Holy Grail ◽  
Viral Therapy

Delivering transgenes to human cells through transduction with viral vectors constitutes one of the most encouraging approaches in gene therapy. Lentivirus-derived vectors are among the most promising vectors for these approaches. When the genetic modification of the cell must be performed in vivo, efficient specific transduction of the cell targets of the therapy in the absence of off-targeting constitutes the Holy Grail of gene therapy. For viral therapy, this is largely determined by the characteristics of the surface proteins carried by the vector. In this regard, an important property of lentiviral vectors is the possibility of being pseudotyped by envelopes of other viruses, widening the panel of proteins with which they can be armed. Here, we discuss how this is achieved at the molecular level and what the properties and the potentialities of the different envelope proteins that can be used for pseudotyping these vectors are.

In Vivo Potency Assay for Adeno-Associated Virus–Based Gene Therapy Vectors Using AAVrh.10 as an Example

2018 ◽  
Vol 29 (3) ◽  
pp. 146-155 ◽  
Author(s):  
Bishnu P. De ◽  
Alvin Chen ◽  
Christiana O. Salami ◽  
Benjamin Van de Graaf ◽  
Jonathan B. Rosenberg ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Potency Assay ◽  
Adeno Associated Virus ◽  
Gene Therapy Vectors

Gene therapy for critical limb ischemia: Per aspera ad astra

2021 ◽  
Vol 21 ◽  
Author(s):  
Vyacheslav Z. Tarantul ◽  
Alexander V. Gavrilenko
Keyword(s):  
Gene Therapy ◽  
Critical Limb Ischemia ◽  
Viral Vectors ◽  
Ex Vivo ◽  
Single Gene ◽  
Public Health Problem ◽  
Limb Ischemia ◽  
Therapeutic Angiogenesis ◽  
Effective Treatment Option

: Peripheral artery diseases remain a serious public health problem. Although there are many traditional methods for their treatment using conservative therapeutic techniques and surgery, gene therapy is an alternative and potentially more effective treatment option especially for “no option” patients. This review treats the results of many years of research and application of gene therapy as an example of treatment of patients with critical limb ischemia. Data on successful and unsuccessful attempts to use this technology for treating this disease are presented. Trends in changing the paradigm of approaches to therapeutic angiogenesis are noted: from viral vectors to non-viral vectors, from gene transfer to the whole organism to targeted transfer to cells and tissues, from single gene use to combination of genes; from DNA therapy to RNA therapy, from in vivo therapy to ex vivo therapy.

scholarly journals Evading the AAV Immune Response in Mucopolysaccharidoses

2020 ◽  
Vol 21 (10) ◽  
pp. 3433
Author(s):  
Matthew Piechnik ◽  
Kazuki Sawamoto ◽  
Hidenori Ohnishi ◽  
Norio Kawamoto ◽  
Yasuhiko Ago ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Immune Response ◽  
Humoral Immune Response ◽  
Transgene Expression ◽  
Evidence Based ◽  
Adeno Associated Virus ◽  
Gene Therapies ◽  
Humoral Immune ◽  
Significant Challenge ◽  
Racial Background

The humoral immune response elicited by adeno-associated virus (AAV)-mediated gene therapy for the treatment of mucopolysaccharidoses (MPS) poses a significant challenge to achieving therapeutic levels of transgene expression. Antibodies targeting the AAV capsid as well as the transgene product diminish the production of glycosaminoglycan (GAG)-degrading enzymes essential for the treatment of MPS. Patients who have antibodies against AAV capsid increase in number with age, serotype, and racial background and are excluded from the clinical trials at present. In addition, patients who have undergone AAV gene therapy are often excluded from the additional AAV gene therapy with the same serotype, since their acquired immune response (antibody) against AAV will limit further efficacy of treatment. Several methods are being developed to overcome this immune response, such as novel serotype design, antibody reduction by plasmapheresis and immunosuppression, and antibody evasion using empty capsids and enveloped AAV vectors. In this review, we examine the mechanisms of the anti-AAV humoral immune response and evaluate the strengths and weaknesses of current evasion strategies in order to provide an evidence-based recommendation on evading the immune response for future AAV-mediated gene therapies for MPS.

Gene Therapy: Paving New Roads in the Treatment of Hemophilia

2019 ◽  
Vol 45 (07) ◽  
pp. 743-750 ◽  
Author(s):  
Gabriela G. Yamaguti-Hayakawa ◽  
Margareth C. Ozelo
Keyword(s):  
Gene Therapy ◽  
Animal Models ◽  
Pediatric Patients ◽  
Viral Vector ◽  
Monogenic Disease ◽  
Adeno Associated Virus ◽  
Underlying Liver Disease ◽  
Experimental Animal Models ◽  
The Road ◽  
Gene Therapy Application

AbstractHemophilia is a monogenic disease with robust clinicolaboratory correlations of severity. These attributes coupled with the availability of experimental animal models have made it an attractive model for gene therapy. The road from animal models to human clinical studies has heralded significant successes, but major issues concerning a previous immunity against adeno-associated virus and transgene optimization remain to be fully resolved. Despite significant advances in gene therapy application, many questions remain pertaining to its use in specific populations such as those with factor inhibitors, those with underlying liver disease, and pediatric patients. Here, the authors provide an update on viral vector and transgene improvements, review the results of recently published gene therapy clinical trials for hemophilia, and discuss the main challenges facing investigators in the field.

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