Specificity of ribozyme designed for mutated DHFR mRNA.
When MOLT-3 human acute leukemia cells were exposed sequentially to trimetrexate (TMQ) and then to methotrexate (MTX), the cells became resistant to antifolate. We designated this subline MOLT-3/TMQ800-MTX10,000. This cell line was found to contain two point mutations in the dihydrofolate reductase (DHFR) gene: a T-->C transition at nucleotide 95 in codon 31, and a T-->A transition at nucleotide 100 in codon 33. In an attempt to specifically inhibit these double-mutated cells, we synthesized a ribozyme which perfectly base-paired with the double-mutated DHFR mRNA. We found that the ribozyme for the double-mutated DHFR mRNA not only cleaved the mutated DHFR RNA, but also efficiently cleaved the wild-type RNA substrate. This observation suggests proceeding with caution when using a ribozyme against a mutated mRNA of an essential enzyme as a specific means of treatment.
Kobayashi
H
H
Derald H. Ruttenberg Cancer Center, Mount Sinai School of Medicine, New York, NY 10029.
Kim
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N
Halatsch
M E
ME
Ohnuma
T
T
D001483:Base Sequence; D002460:Cell Line; D006801:Humans; D008727:Methotrexate / Q000494:pharmacology; D008969:Molecular Sequence Data; D017354:Point Mutation; D016337:RNA, Catalytic / Q000138:chemical synthesis / Q000235:genetics* / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism*; D013379:Substrate Specificity; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*; D016597:Trimetrexate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D016337:RNA, Catalytic; D012333:RNA, Messenger; D013762:Tetrahydrofolate Dehydrogenase; D016597:Trimetrexate; D008727:Methotrexate