Inhibition of C-raf expression by antisense oligonucleotides extends heart allograft survival in rats.
BACKGROUND
C-raf is a well-characterized serine/ threonine (Ser/Thr) protein kinase that is involved in the transduction of multiple signals of T cells. We demonstrate that the inhibition of C-raf mRNA expression prolongs heart 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 positions 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 five 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 expression in two cell lines. Similarly, each of the ME C-raf antisense oligos produced better heart allograft survival rates than did PS C-raf oligo. Furthermore, although the combination of PS C-raf antisense oligo with sirolimus (SRL) acted synergistically to extend heart allograft survival, the effect was potentiated by either of the ME-modified oligos.


CONCLUSIONS
C-raf inhibition extends heart allograft survival, and ME-modification potentiates antisense activity.
Stepkowski
S M
SM
Division of Immunology and Organ Transplantation, The University of Texas Medical School at Houston, 77030, USA.
Qu
X
X
Wang
M E
ME
Tian
L
L
Chen
W
W
Wancewicz
E V
EV
Johnston
J F
JF
Bennett
C F
CF
Monia
B P
BP
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D005786:Gene Expression Regulation / Q000187:drug effects*; D006085:Graft Survival / Q000187:drug effects / Q000235:genetics*; D016027:Heart Transplantation / Q000276:immunology / Q000502:physiology*; D006801:Humans; D051379:Mice; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology*; D019908:Proto-Oncogene Proteins c-raf / Q000235:genetics*; D011519:Proto-Oncogenes; D051381:Rats; D011912:Rats, Inbred ACI; D011917:Rats, Inbred Lew; D013873:Thionucleotides; D014184:Transplantation, Homologous
D020319:Oligodeoxyribonucleotides, Antisense; D013873:Thionucleotides; D019908:Proto-Oncogene Proteins c-raf