title: Spliceosome-mediated RNA trans-splicing facilitates targeted delivery of suicide genes to cancer cells.
abstract: Patients suffering from recessive dystrophic epidermolysis bullosa (RDEB), a hereditary blistering disease of epithelia, show susceptibility to develop highly aggressive squamous cell carcinoma (SCC). Tumors metastasize early and are associated with mortality in the 30th-40th years of life in this patient group. So far, no adequate therapy is available for RDEB SCC. An approach is suicide gene therapy, in which a cell death-inducing agent is introduced to cancer cells. However, lack of specificity has constrained clinical application of this modality. Therefore, we used spliceosome-mediated RNA trans-splicing technology, capable of replacing a tumor-specific transcript with one encoding a cell death-inducing peptide/toxin, to provide tumor-restricted expression. We designed 3' pre-trans-splicing molecules (PTM) and evaluated their efficiency to trans-splice an RDEB SCC-associated target gene, the matrix metalloproteinase-9 (MMP9), in a fluorescence-based test system. A highly efficient PTM was further adapted to insert the toxin streptolysin O (SLO) of Streptococcus pyogenes into the MMP9 gene. Transfection of RDEB SCC cells with the SLO-PTM resulted in cell death and induction of toxin function restricted to RDEB SCC cells. Thus, RNA trans-splicing is a suicide gene therapy approach with increased specificity to treat highly malignant SCC tumors.
authors.affiliation: Division of Molecular Dermatology and EB House Austria, Department of Dermatology, Paracelsus Medical University, Salzburg, Muellner Hauptstrasse 48, 5020 Salzburg, Austria. c.gruber@salk.at
authors.forename: Christina
authors.initials: C
authors.lastname: Gruber
authors.forename: Iris K
authors.initials: IK
authors.lastname: Gratz
authors.forename: Eva M
authors.initials: EM
authors.lastname: Murauer
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Mayr
authors.forename: Ulrich
authors.initials: U
authors.lastname: Koller
authors.forename: Leena
authors.initials: L
authors.lastname: Bruckner-Tuderman
authors.forename: Guerrino
authors.initials: G
authors.lastname: Meneguzzi
authors.forename: Helmut
authors.initials: H
authors.lastname: Hintner
authors.forename: Johann W
authors.initials: JW
authors.lastname: Bauer
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002294:Carcinoma, Squamous Cell / Q000209:etiology* / Q000235:genetics* / Q000628:therapy; D002460:Cell Line; D002470:Cell Survival / Q000235:genetics; D016108:Epidermolysis Bullosa Dystrophica / Q000150:complications* / Q000235:genetics* / Q000628:therapy; D005434:Flow Cytometry; D023061:Gene Order; D018014:Gene Transfer Techniques; D041722:Genes, Transgenic, Suicide / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D020780:Matrix Metalloproteinase 9 / Q000235:genetics* / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D017381:Spliceosomes / Q000378:metabolism*; D013301:Streptolysins / Q000235:genetics* / Q000378:metabolism; D020040:Trans-Splicing / Q000235:genetics*
chemical_list: D001426:Bacterial Proteins; D013301:Streptolysins; C025964:streptolysin O; D020780:Matrix Metalloproteinase 9