The antisense c-Ha-ras oligodeoxynucleotides inhibit the proliferation of the gastrocarcinoma-DNA-transformed rat 3-3 cells and reduce the p21 expression of the cells.
The Rat 3-3 is a secondary transformant of the rat fibroblast cell line (Rat-1) transfected with total DNA of a gastrocarcinoma cell line BGC-823. The cells over-express the c-Ha-ras oncogene which contains point mutation at the 12th codon. In order to determine how the activated c-Ha-ras oncogene expression governs the cell's transformation, two pentadecadeoxy-nucleotides AS-1 and AS-2 were synthesized. AS-1 was complementary to the single strand of the first three codons and the upstream sequence close to the ribosome binding site of c-Ha-ras mRNA. AS-2 was complementary to the 3' end of the first intron and the 5' end of the second exon of c-Ha-ras unripe RNA enclosed in the nucleus. The oligonucleotides could block either the translation of c-Ha-ras mRNA or the splicing of c-Ha-ras unripe RNA, thus inhibiting the expression of the activated c-Ha-ras oncogene and the proliferation of the transformed cells Rat 3-3. The inhibitory effect increased with a growing concentration of the antisense oligodeoxynucleotide (from 4-10 mumol/L) and reached its peak at 12 h after Rat 3-3 was treated with AS-1 and AS-2. This effect became weaker afterwards. The p21 level, product of c-Ha-ras, in the Rat 3-3, was examined by ELISA method. The result shows that the amount of p21 in the growth-inhibited cells is about 30% of that of the control cells.
Xu
Y
Y
Biochemistry Department, Beijing Institute for Cancer Research, PRC.
Deng
G R
GR
Liang
Y Y
YY
Li
C
C
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D004273:DNA, Neoplasm / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000166:cytology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D011905:Genes, ras; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D014176:Protein Biosynthesis / Q000187:drug effects; D016283:Proto-Oncogene Proteins p21(ras) / Q000032:analysis*; D012326:RNA Splicing / Q000187:drug effects; D051381:Rats; D013274:Stomach Neoplasms / Q000235:genetics / Q000473:pathology*; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000187:drug effects
D004273:DNA, Neoplasm; D016376:Oligonucleotides, Antisense; D016283:Proto-Oncogene Proteins p21(ras)