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Development of a stable Sf9 insect cell line to produce VSV-G pseudotyped baculoviruses

Gene Therapy volume 31, pages 187–194 (2024)Cite this article

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Abstract

Baculoviruses have shown great potential as gene delivery vectors in mammals, although their effectiveness in transferring genes varies across different cell lines. A widely employed strategy to improve transduction efficiency is the pseudotyping of viral vectors. In this study, we aimed to develop a stable Sf9 insect cell line that inducibly expresses the G-protein of the vesicular stomatitis virus to pseudotype budded baculoviruses. It was obtained by inserting the VSV-G gene under the control of the very strong and infection-inducible pXXL promoter and was subsequently diluted to establish oligoclonal lines, which were selected by the fusogenic properties of VSV-G and its expression levels in infected cells and purified budded virions. Next, to enhance the performance of the cell line, the infection conditions under which functional pseudotyped baculoviruses are obtained were optimized. Finally, different baculoviruses were pseudotyped and the expression of the transgene was quantified in mammalian cells of diverse origins using flow cytometry. The transduction efficiency of pseudotyped baculovirus consistently increased across all tested mammalian cell lines compared with control viruses. These findings demonstrate the feasibility and advantages of improving gene delivery performance without the need to insert the pseudotyping gene into the baculoviral genomes.

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Fig. 1: Schematic representation of the obtention and characterization of Sf9 VSV-G cell line.
Fig. 2: Evaluation of VSV-G expression and functionality in the sublines and clones.
Fig. 3: Transduction efficiency of pseudotyped baculovirus obtained under different experimental conditions.
Fig. 4: Transduction efficiencies of pseudotyped baculoviruses.

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Data availability

The datasets generated during the current study are available from the corresponding author on reasonable request.

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Acknowledgements

MGL, OT and VA are researchers from CONICET. MPP has a fellowship from ANPCyT.

Funding

This work was supported by Fondo Nacional de Ciencia y Tecnología (FONCyT), grant numbers PICT 3891 and PICT 0268, CONICET, PIP 592 and INTA, 2019-PD-E5-I105-001.

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Author notes

  1. These authors contributed equally: María del Pilar Plastine, Sabrina Amalfi.

Authors and Affiliations

  1. Instituto de Agrobiotecnología y Biología Molecular (IABIMO), Instituto Nacional de Tecnología Agropecuaria (INTA), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), De los Reseros y N. Repetto S/N, Hurlingham, B1686IGC, Buenos Aires, Argentina

    María del Pilar Plastine, Sabrina Amalfi, María Gabriela López, María José Gravisaco, Oscar Taboga & Victoria Alfonso

  2. Universidad Nacional de Hurlingham, Instituto de Biotecnología, Av. Vergara 2222, Villa Tesei, Hurlingham, B1688GEZ, Buenos Aires, Argentina

    Sabrina Amalfi

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  1. María del Pilar Plastine
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  3. María Gabriela López
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Contributions

Conceptualization, OT and VA; Methodology, MJG, MGL and VA; Investigation, MPP and SA; Writing – Original Draft, MPP, SA and VA; Writing – Review & Editing, MGL, OT and VA; Funding Acquisition, OT and VA; Supervision, VA.

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Correspondence to Oscar Taboga or Victoria Alfonso.

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Plastine, M.d.P., Amalfi, S., López, M.G. et al. Development of a stable Sf9 insect cell line to produce VSV-G pseudotyped baculoviruses. Gene Ther 31, 187–194 (2024). https://doi.org/10.1038/s41434-024-00442-4

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