Despite scientific discoveries in the field of gene and cell therapy, some diseases still have no effective treatment. Advances in genetic engineering methods have enabled the development of effective gene therapy methods for various diseases based on adeno-associated viruses (AAVs). Today, many AAV-based gene therapy medications are being investigated in preclinical and clinical trials, and new ones are appearing on the market. In this article, we present a review of AAV discovery, properties, different serotypes, and tropism, and a following detailed explanation of their uses in gene therapy for disease of different organs and systems.

Sphingolipidoses are defined as a group of rare hereditary diseases resulting from mutations in the genes encoding lysosomal enzymes. This group of lysosomal storage diseases includes more than 10 genetic disorders, including GM1-gangliosidosis, Tay–Sachs disease, Sandhoff disease, the AB variant of GM2-gangliosidosis, Fabry disease, Gaucher disease, metachromatic leukodystrophy, Krabbe disease, Niemann–Pick disease, Farber disease, etc. Enzyme deficiency results in accumulation of sphingolipids in various cell types, and the nervous system is also usually affected. There are currently no known effective methods for the treatment of sphingolipidoses; however, gene therapy seems to be a promising therapeutic variant for this group of diseases. In this review, we discuss gene therapy approaches for sphingolipidoses that are currently being investigated in clinical trials, among which adeno-associated viral vector-based approaches and transplantation of hematopoietic stem cells genetically modified with lentiviral vectors seem to be the most effective.

Passaged cell lines represent currently an integral component in various studies of malignant neoplasms. These cell lines are utilized for drug screening both in monolayer cultures or as part of three-dimensional (3D) tumor models. They can also be used to model the tumor microenvironment in vitro and in vivo through xenotransplantation into immunocompromised animals. However, immortalized cell lines have some limitations of their own. The homogeneity of cell line populations and the extensive passaging in monolayer systems make these models distant from the original disease. Recently, there has been a growing interest among scientists in the use of primary cell lines, as these are passaged directly from human tumor tissues. In this case, cells retain the morphological and functional characteristics of the tissue from which they were derived, an advantage often not observed in passaged cultures. This review highlights the advantages and limitations of passaged and primary cell cultures, their similarities and differences, as well as existing test systems that are based on primary and passaged cell cultures for drug screening purposes.

Currently, the development of adeno-associated virus (AAV)-based gene therapy is a promising method for treating various diseases and is gaining increasing popularity. However, the use of AAV has certain drawbacks and faces limitations such as immune responses and an increased risk of insertional mutagenesis, which have not always been adequately considered in the context of AAV therapy. Moreover, a significant limitation for the application of AAV lies in the challenge of producing it in large quantities. This article discusses the use of AAV in treating various diseases, reviews AAV production approaches, highlights challenges with insufficient viral titers during production, and explores potential solutions at key stages of AAV drug production.

Metachromatic leukodystrophy (MLD) is a hereditary neurodegenerative disease characterized by demyelination and motor and cognitive impairments due to deficiencies of the lysosomal enzyme arylsulfatase A (ARSA) or the saposin B activator protein (SapB). Current treatments are limited; however, gene therapy using adeno-associated virus (AAV) vectors for ARSA delivery has shown promising results. The main challenges for MLD gene therapy include optimizing the AAV dosage, selecting the most effective serotype, and determining the best route of administration for ARSA delivery into the central nervous system. This study aims to evaluate the safety and efficacy of AAV serotype 9 encoding ARSA (AAV9-ARSA) gene therapy when administered intravenously or intrathecally in minipigs, a large animal model with anatomical and physiological similarities to humans. By comparing these two administration methods, this study contributes to the understanding of how to improve the effectiveness of MLD gene therapy and offers valuable insights for future clinical applications.

Neutralizing antibodies (nAbs) are an important component of the immune system, which plays a dual role in modern medicine. On the one hand, they significantly limit the effectiveness of gene therapy based on viral vectors, reducing the effectiveness of treatment of diseases such as spinal muscular atrophy, which is especially evident with repeated administration of therapeutic vectors. On the other hand, nAbs is a promising tool for combating viral infections. This review systematizes current data on the mechanisms of nAbs formation against AAV vectors, analyzes the factors influencing their production, and discusses strategies to overcome this limitation, including modification of vectors and the development of methods to suppress the immune response. Special attention is paid to the prospects of using nAbs as therapeutic agents against viral infections. The key problems and possible directions of research development in this area are considered, which is important for improving approaches to the treatment of both rare genetic and infectious diseases.