Current Gene Therapy - Volume 18, Issue 3, 2018

Small activating RNAs (saRNAs) are small double-stranded RNAs that could mediate the target-specific gene expression by targeting selected sequences in gene promoters at both the transcriptional and epigenetic levels. This phenomenon of gene manipulation is known as RNA activation (RNAa), which opens up a new pathway for RNA-based gene therapeutics in contrast to RNA interference. Although the exact molecular mechanism of RNAa mediated by saRNAs still remains foggy, some studies have provided the possible ones to explain it. Furthermore, mounting evidence exhibit that saRNAs not only provide a new approach to study gene function and manipulate transcriptional activity, but also promise a great potential for clinical therapy against various diseases, especially cancer. Cancer-associated genes could be up-regulated by saRNAs to modulate cell cycle and proliferation, induce cell senescence and apoptosis, inhibit cancer cell invasion and migration, and reverse chemotherapy resistance. Herein, we summarize the known mechanisms of saRNAs on up-regulating specific gene expression and focus on the potential applications of saRNAs in gene therapy. In addition, some concerns about mechanisms and challenges for delivery of saRNAs are involved in this review. The precise mechanisms of saRNAs need to be further illustrated and some novel delivery systems for saRNAs are expected to be developed for clinical applications.

The present review examines whether the microRNA 7 (miR-7) holds potential for slowing Parkinson's disease (PD) progression. First, the accurate expression of miR-7 allows for normal development, physiology, and neurogenesis in the central nervous system, also keeping alpha-synuclein (α-Syn) at the physiological level. Second, patients with PD and parkinsonian MPTP-induced animals exhibit a significant decrease of miR-7 in brain areas associated with dopaminergic neurodegeneration. Depletion of miR-7 in the substantia nigra of clinical samples is related to α-Syn accumulation, loss of dopaminergic cells, and reduction of dopamine in the striatum. Therefore, the goal of a miR-7- replacement therapy is to downregulate α-Syn and other PD-related genes, achieving multi-target benefits regarding oxidative stress, mitochondrial health, cell glycolysis, apoptosis, and inhibition of inflammasome activation. While a disease-modifying drug is a major unmet need for the clinical management of PD, an miR-7-replacement therapy presents a striking potential against critical mechanisms of neuropathology. Such innovative treatment would reduce α-Syn accumulation in the Lewy bodies and preserve remaining neurons yet viable at the time of diagnosis, thus slowing disease progression from the early phase of PD characterized by a relatively mild motor impairment to an advanced and more disabling stage.

Musculoskeletal conditions are a major public health problem. Approximately 66 million individuals seek medical attention for a musculoskeletal injury in the United States, with current medical costs being estimated at $873 billion annually. Despite advances in pharmaceuticals, implant materials and surgical techniques, there remains an unmet clinical need for successful treatment of challenging musculoskeletal injuries and pathologic conditions, particularly in the setting of compromised biological environments. Tissue engineering via gene therapy attempts to provide an alternative treatment strategy to address the deficits associated with conventional approaches. The transfer of specific target genes coding for proteins with therapeutic or regenerative properties to target cells and tissues in the disease environment allows for their sustained production and release specifically at the site of interest. The increasing reports of success with gene therapy-based treatments in the clinical management of a variety of diseases provide genuine optimism that similar methods can be adapted for mainstream clinical application in musculoskeletal disorders. In preclinical studies, gene therapy has been successfully used to treat cartilaginous, bone, skeletal muscle, tendon, ligament and intervertebral disk injuries. In addition, gene therapy is being assessed in clinical trials for its safety and therapeutic potential in osteoarthritis. This review will specifically address the clinical potential, preclinical data and future hurdles for gene therapy to be a viable clinical entity for the treatment of fracture nonunion and difficult bone repair scenarios, articular cartilage repair and osteoarthritis.

Background: Following joint trauma, a posttraumatic inflammatory cascade drives degeneration of the joint. We aimed to assess whether transduction of chondrocytes with AAV5 overexpressing the immunomodulatory cytokine IL-10 would have protective effects in pellet cultures stimulated with IL-1β. Methods: Chondrocytes were isolated from 3 healthy horses and were transduced with AAV5-IL-10 at a dose of 1 x 105vg/cell. Chondrocyte pellets were formed by centrifugation and were stimulated with IL-1β starting 48 hours following transduction. After 2, 6 and 14 days in culture, supernatants were collected for cytokine analysis and RNA was isolated from cells for gene expression analysis. Pellets were also collected for biochemical analysis. Results: Transduction of chondrocytes led to significant increases in IL-10 expression. IL-10 expression was further enhanced by IL-1β stimulation. IL-10 overexpression led to significantly decreased expression of IL-1β and ADAMTS4. PGE2 synthesis was also significantly decreased. IL-1β mediated suppression of GAG synthesis was not rescued by IL-10. Conclusions: Overexpression of IL-10 modulates the inflammatory response in chondrocytes, which may mitigate some of the deleterious effects of pro-inflammatory cascades in the posttraumatic joint. AAV5-IL-10 led to efficient and sustained overexpression of IL-10 in chondrocytes and could represent a viable treatment option for preventing osteoarthritis.

Background: The West Nile Virus (WNV) has emerged as one of the most significant arboviral infection in many parts of the world and is associated with the encephalitis affecting mainly human and horses. In spite of the fact that the WNV is threat for the public health, there is no vaccine or therapeutic available for the treatment of WNV. Methods: In this study, we tested a novel RNA interference based technique to inhibit WNV replication in Vero cells. Two siRNAs were designed against the NS2A and NS5 regions of WNV which are highly conserved among Flaviviruses as it play important role in apoptosis and in viral replication respectively. In addition to this, dual functional siRNA is designed by joining an immunostimulatroy motif with the NS2A and NS5 specific siRNA. The antiviral activity was evaluated by detecting both the infectious virus and its genome. Results: The bifunctional siRNA resulted in significant reduction of virus titre in siRNA transfected cells as compared to controls. The antiviral efficacy was most effective at 48hr post infection. These results were in accordance with the quantitative RT-PCR assay revealing similar reduction in WNV genomic RNA. The expression of housekeeping gene was not affected by the siRNA indicating no off target effect and non-interference in cellular mechanism. Conclusion: Thus, this bifunctional siRNA intervention paves the new way for therapeutic treatment of WNV disease.