Prolonged activity of the transposase helper may raise safety concerns during DNA transposon-based gene therapy

Imre, Gergely and Takács, Bertalan Vilmos and Czipa, Erik and Bakné Drubi, Andrea and Jaksa, Gábor and Latinovics, Dóra and Nagy, Andrea and Karkas, Réka and Hudoba, Liza and Vásárhelyi, Bálint Márk and Pankotai-Bodó, Gabriella and Blastyák, András and Hegedűs, Zoltán and Germán, Péter and Bálint, Balázs and Abdullah, Khaldoon Sadiq Ahmed and Kopasz, Anna Georgina and Kovács, Anita Kármen and Nagy, László and Sükösd, Farkas and Pintér, Lajos and Rülicke, Thomas and Barta, Endre and Nagy, István and Haracska, Lajos and Mátés, Lajos (2023) Prolonged activity of the transposase helper may raise safety concerns during DNA transposon-based gene therapy. MOLECULAR THERAPY-METHODS AND CLINICAL DEVELOPMENT, 29. pp. 145-159. ISSN 2329-0501

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Abstract

DNA transposon-based gene delivery vectors represent a promising new branch of randomly integrating vector development for gene therapy. For the side-by-side evaluation of the piggyBac and Sleeping Beauty systems—the only DNA transposons currently employed in clinical trials—during therapeutic intervention, we treated the mouse model of tyrosinemia type I with liver-targeted gene delivery using both transposon vectors. For genome-wide mapping of transposon insertion sites we developed a new next-generation sequencing procedure called streptavidin-based enrichment sequencing, which allowed us to identify approximately one million integration sites for both systems. We revealed that a high proportion of piggyBac integrations are clustered in hot regions and found that they are frequently recurring at the same genomic positions among treated animals, indicating that the genome-wide distribution of Sleeping Beauty-generated integrations is closer to random. We also revealed that the piggyBac transposase protein exhibits prolonged activity, which predicts the risk of oncogenesis by generating chromosomal double-strand breaks. Safety concerns associated with prolonged transpositional activity draw attention to the importance of squeezing the active state of the transposase enzymes into a narrower time window.

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