Inhibition of C-raf expression by antisense oligonucleotides extends heartallograft survival in rats

Background. C-raf is a well-characterized serine/threonine (Ser/Thr) protei n kinase that is involved in the transduction of multiple signals of T cell s. We demonstrate that the inhibition of C-raf mRNA expression prolongs hea rt allograft survival. Methods. Three 20-mer C-raf antisense oligonucleotides, each with identical sequences, were synthesized with different chemical modifications: one as a uniform phosphorothioate oligodeoxynucleotide (PS oligo), a second with a PS backbone and 2'-methoxyethyl (ME) substitutions at the 2'-sugar positio ns in the first and last five nucleotides, and a third with a mixed PS and phosphodiester (PD) backbone and ME modifications on the first and last fiv e nucleotides, Results. Both ME-modified C-raf antisense oligos were at least 5-fold more effective than the PS C-raf antisense oligo in blocking C-raf mRNA expressi on in two cell lines. Similarly, each of the ME C-raf antisense oligos prod uced better heart allograft survival rates than did PS C-raf oligo, Further more, although the combination of PS C-raf antisense oligo with sirolimus ( SRL) acted synergistically to extend heart allograft survival, the effect w as potentiated by either of the ME-modified oligos, Conclusions. C-raf inhibition extends heart allograft survival, and ME-modi fication potentiates antisense activity.