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Background <p>Neurological disorders are the second leading cause of death and the leading cause of disability in the world<b>.</b> Thus, the development of novel disease-modifying strategies is clearly warranted. We have previously developed a therapeutic approach using mouse targeted rabies virus glycoprotein (RVG) extracellular vesicles (EVs) to deliver minicircles (MCs) expressing shRNA (shRNA-MCs) to induce long-term α-synuclein down-regulation. Although the previous therapy successfully reduced the pathology, the clinical translation was extremely unlikely since they were mouse extracellular vesicles.</p> Methods <p>To overcome this limitation, we developed a source of human RVG-EVs compatible with a personalized therapy using immature dendritic cells. Human peripheral blood monocytes were differentiated in vitro into immature dendritic cells, which were transfected to express the RVG peptide. RVG-EVs containing shRNA-MCs, loaded by electroporation, were injected intravenously in the α-synuclein performed fibril (PFF) mouse model. Level of α-synuclein, phosphorylated α-synuclein aggregates, dopaminergic neurons and motor function were evaluated 90&#xa0;days after the treatment. To confirm that EVs derived from patients were suitable as a vehicle, proteomic analysis of EVs derived from control, initial and advanced Parkinson’s disease was performed.</p> Results <p>The shRNA-MCs could be successfully loaded into human RVG-EVs and downregulate α-synuclein in SH-SY5Y cells. Intravenous injection of the shRNA-MC-loaded RVG-EVs induced long-term downregulation of α-synuclein mRNA expression and protein level, decreased α-synuclein aggregates, prevented dopaminergic cell death and ameliorated motor impairment in the α-synuclein PFF mouse model. Moreover, we confirmed that the EVs from PD patients are suitable as a personalized therapeutic vehicle.</p> Conclusion <p>Our study confirmed the therapeutic potential of shRNA-MCs delivered by human RVG-EVs for long-term treatment of neurodegenerative diseases. These results pave the way for clinical use of this approach.</p>

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Development of human targeted extracellular vesicles loaded with shRNA minicircles to prevent parkinsonian pathology

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年5月26日

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  • Maria Izco,
  • Carlos Sola,
  • Martin Schleef,
  • Marco Schmeer,
  • María de Toro,
  • Guglielmo Verona,
  • Estefania Carlos,
  • Alejandro Reinares-Sebastian,
  • Sandra Colina,
  • Maria Eugenia Marzo-Sola,
  • Josune Garcia-Sanmartin,
  • Joaquín Fernández-Irigoyen,
  • Enrique Santamaría,
  • Rodolfo Mugica-Vidal,
  • Javier Blesa,
  • Lydia Alvarez-Erviti

摘要

Background

Neurological disorders are the second leading cause of death and the leading cause of disability in the world. Thus, the development of novel disease-modifying strategies is clearly warranted. We have previously developed a therapeutic approach using mouse targeted rabies virus glycoprotein (RVG) extracellular vesicles (EVs) to deliver minicircles (MCs) expressing shRNA (shRNA-MCs) to induce long-term α-synuclein down-regulation. Although the previous therapy successfully reduced the pathology, the clinical translation was extremely unlikely since they were mouse extracellular vesicles.

Methods

To overcome this limitation, we developed a source of human RVG-EVs compatible with a personalized therapy using immature dendritic cells. Human peripheral blood monocytes were differentiated in vitro into immature dendritic cells, which were transfected to express the RVG peptide. RVG-EVs containing shRNA-MCs, loaded by electroporation, were injected intravenously in the α-synuclein performed fibril (PFF) mouse model. Level of α-synuclein, phosphorylated α-synuclein aggregates, dopaminergic neurons and motor function were evaluated 90 days after the treatment. To confirm that EVs derived from patients were suitable as a vehicle, proteomic analysis of EVs derived from control, initial and advanced Parkinson’s disease was performed.

Results

The shRNA-MCs could be successfully loaded into human RVG-EVs and downregulate α-synuclein in SH-SY5Y cells. Intravenous injection of the shRNA-MC-loaded RVG-EVs induced long-term downregulation of α-synuclein mRNA expression and protein level, decreased α-synuclein aggregates, prevented dopaminergic cell death and ameliorated motor impairment in the α-synuclein PFF mouse model. Moreover, we confirmed that the EVs from PD patients are suitable as a personalized therapeutic vehicle.

Conclusion

Our study confirmed the therapeutic potential of shRNA-MCs delivered by human RVG-EVs for long-term treatment of neurodegenerative diseases. These results pave the way for clinical use of this approach.