Homologous human and murine antisense oligonucleotides targeting stat6. Functional effects on germline cepsilon transcript.
Interleukin (IL)-4 and (IL)-13 induce immunoglobulin (Ig)E synthesis via activation of the transcription factor signal transducer and activator of transcription (Stat)6. The present study describes the identification and characterization of antisense oligonucleotides to Stat6 as an approach to interrupt IL-4 and IL-13 signaling and thereby to attenuate germline Cepsilon transcription, a prerequisite to IgE synthesis. A limited gene-walk was performed with chemically modified oligonucleotides to identify sequences capable of downregulating both human and murine Stat6. A chimeric oligonucleotide (9b, base sequence GTGAGGTCCTGTTCAGTGGG) demonstrated high levels of antisense activity in both species. Further characterization of 9b showed a dose-dependent Stat6 messenger RNA (mRNA) and protein downregulation (concentration that produces 50% inhibition of effect = 168 and 215 nM, respectively) through a ribonuclease H-dependent antisense mechanism with no effect on closely related members of the Stat family. Further, pretreatment of DND39 cells (human Burkitt lymphoma cell line) with oligonucleotide 9b before IL-4 stimulation successfully downregulated germline Cepsilon transcription. Because Stat6 represents an attractive but technically challenging drug discovery target, antisense oligonucleotides may provide an alternative approach to low molecular-weight compounds for inhibiting IL-4 and IL-13 signaling.
Hill
S
S
Molecular and Cell Biology Unit, Novartis Horsham Research Centre, West Sussex, United Kingdom. sandra.hill@pharma.novartis.com
Herlaar
E
E
Le Cardinal
A
A
van Heeke
G
G
Nicklin
P
P
D000818:Animals; D002460:Cell Line; D015536:Down-Regulation; D005786:Gene Expression Regulation; D006801:Humans; D007073:Immunoglobulin E / Q000096:biosynthesis / Q000235:genetics*; D018793:Interleukin-13 / Q000037:antagonists & inhibitors; D015847:Interleukin-4 / Q000037:antagonists & inhibitors; D051379:Mice; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D016914:Ribonuclease H / Q000378:metabolism; D050800:STAT6 Transcription Factor; D015398:Signal Transduction; D015534:Trans-Activators / Q000096:biosynthesis / Q000235:genetics*; D014158:Transcription, Genetic
D018793:Interleukin-13; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D050800:STAT6 Transcription Factor; C494101:STAT6 protein, human; C494100:Stat6 protein, mouse; D015534:Trans-Activators; D015847:Interleukin-4; D007073:Immunoglobulin E; D016914:Ribonuclease H