Target site search and effective inhibition of leukaemic cell growth by a covalently closed multiple anti-sense oligonucleotide to c-myb.
Systematic secondary structure simulation of a target mRNA sequence is shown to be effective for locating a good anti-sense target site. Multiple selected anti-sense sequences were placed in a single molecule. The anti-sense oligonucleotide (oligo) was covalently closed to avoid exonuclease activities and was designated CMAS (covalently closed multiple anti-sense)-oligo. CMAS-oligo was found to be stable, largely preserving its structural integrity after 24 h of incubation in the presence of either exonuclease III or serum. When human c-myb mRNA was targeted by the c-myb CMAS-oligo, expression of the gene was completely abolished. Further, tumour cell growth was inhibited by 82+/-3% as determined by an MTT [3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay and by 90+/-1% by [(3)H]thymidine incorporation. When a leukaemic cell line K562 was treated with CMAS-oligo, colony formation on soft agarose was also decreased by 93%. In contrast, treatment with a scrambled control oligo did not significantly inhibit leukaemic cell growth. These results suggest that a rational target site search is possible for an anti-sense oligo and that CMAS-oligo can be employed as an effective anti-sense agent with enhanced stability.
Moon
I J
IJ
Institute for Medical Science, Dongsan Medical Center, Keimyung University, Dongsandong 194, Chunggu, Taegu 700-310, Korea.
Lee
Y
Y
Kwak
C S
CS
Lee
J H
JH
Choi
K
K
Schreiber
A D
AD
Park
J G
JG
D002455:Cell Division / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000235:genetics*; D018390:Gene Targeting; D020570:Genes, myb; D006801:Humans; D007938:Leukemia / Q000235:genetics* / Q000473:pathology*; D016376:Oligonucleotides, Antisense / Q000235:genetics* / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics; D014407:Tumor Cells, Cultured
D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger