Correction of DNA protein kinase deficiency by spliceosome-mediated RNA trans-splicing and sleeping beauty transposon delivery

Hatem Zayed, Lily Xia, Anton Yerich, Stephen R. Yant, Mark A. Kay, M. Puttaraju, Gerard J. McGarrity, David L. Wiest, R. Scott McIvor, Jakub Tolar, Bruce R. Blazar

Research output: Contribution to journalArticlepeer-review

18 Scopus citations


Spliceosome-mediated RNA trans-splicing (SMaRT) is an emerging technology for the repair of defective pre-messenger RNA (pre-mRNA) molecules. It is especially useful in the treatment of genetic disorders involving large genes. Although viral vectors have been used for achieving long-lasting expression of trans-splicing molecules, the immunogenicity and suboptimal safety profiles associated with viral-based components could limit the widespread application of SMaRT in the repair of genetic defects. Here, we tested whether the non-viral Sleeping Beauty (SB) transposon system could mediate stable delivery of trans-splicing molecules designed to correct the genetic defect responsible for severe combined immune deficiency (SCID). This immunological disorder is caused by a point mutation within the 12.4 kilobase (kb) gene encoding the DNA protein kinase catalytic subunit (DNA-PKcs) and is associated with aberrant DNA repair, defective T- and B-cell production, and hypersensitivity to radiation-induced injury. Using a novel SB-based trans-splicing vector, we demonstrate stable mRNA correction, proper DNA-PKcs protein production, and conference of a radiation-resistant phenotype in a T-cell thymoma cell line and SCID multipotent adult progenitor cells (MAPCs). These results suggest that SB-based trans-splicing vectors should prove useful in facilitating the correction of endogenous mutated mRNA transcripts, including the DNA-PKcs defect present in SCID cells.

Original languageEnglish (US)
Pages (from-to)1273-1279
Number of pages7
JournalMolecular Therapy
Issue number7
StatePublished - Jul 2007

Bibliographical note

Funding Information:
This work was supported by an NIH grant NIH R01 HL52952.

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