title: Antisense-mediated exon skipping to correct IL-12Rbeta1 deficiency in T cells.
abstract: Patients with Mendelian susceptibility to mycobacterial disease have severe, recurrent life-threatening infections with otherwise poorly pathogenic mycobacteria and salmonellae. The extreme susceptibility is the result of genetic defects in the interleukin-12/interferon-gamma (IL-12/IFN-gamma) pathway. The infections are difficult to treat, and therapeutic options are limited. We explored the feasibility of antisense-mediated exon skipping as therapy for Mendelian susceptibility to mycobacterial disease with cells from a complete IL-12Rbeta1(-/-) patient. Expression constructs were first studied to determine whether IL12RB1 lacking exon 2 encodes a functional protein. The IL-12Rbeta1 expression construct lacking exon 2 was expressed on T cells. On IL-12 or IL-23 stimulation, this construct phosphorylated similar amounts of STAT1, STAT3, and STAT4 and induced similar amounts of IFN-gamma compared with a normal IL-12Rbeta1 construct. Antisense oligonucleotides (AONs) directed at exon 2 resulted in transcripts lacking exon 2 in both controls' and patients' T cells. In IL-12Rbeta1(-/-) cells, skipping of exon 2 led to expression of IL-12Rbeta1 on the cell surface and responsiveness to IL-12. We showed that IL12RB1 lacking exon 2 encodes a functional IL-12Rbeta1. We demonstrated that T cells can be highly efficiently transduced with AONs and are amenable to antisense-mediated exon skipping. Furthermore, we showed that exon skipping (partly) corrects the IL-12Rbeta1 deficiency in patients' cells.
authors.affiliation: Department of Infectious Diseases, Leiden University Medical Center, Leiden, The Netherlands. E.van_de_Vosse@LUMC.nl.
authors.forename: Esther
authors.initials: E
authors.lastname: van de Vosse
authors.forename: Els M
authors.initials: EM
authors.lastname: Verhard
authors.forename: Roelof A
authors.initials: RA
authors.lastname: de Paus
authors.forename: Gerard J
authors.initials: GJ
authors.lastname: Platenburg
authors.forename: Judith C T
authors.initials: JC
authors.lastname: van Deutekom
authors.forename: Annemieke
authors.initials: A
authors.lastname: Aartsma-Rus
authors.forename: Jaap T
authors.initials: JT
authors.lastname: van Dissel
mesh_terms: D015153:Blotting, Western; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D005091:Exons / Q000235:genetics*; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D009000:Monocytes / Q000166:cytology / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000235:genetics*; D010766:Phosphorylation; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018123:Receptors, Interleukin / Q000235:genetics / Q000378:metabolism; D053707:Receptors, Interleukin-12 / Q000037:antagonists & inhibitors / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050794:STAT1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050795:STAT2 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism; D013601:T-Lymphocytes / Q000378:metabolism*
chemical_list: C508438:IL12RB1 protein, human; C515500:IL23R protein, human; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D018123:Receptors, Interleukin; D053707:Receptors, Interleukin-12; D050794:STAT1 Transcription Factor; D050795:STAT2 Transcription Factor; C494084:STAT2 protein, human; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; D007371:Interferon-gamma