J. Biol. Chem., Vol. 259, Issue 21, 12964-12969, 11, 1984
Neplanocin A. A cyclopentenyl analog of adenosine with specificity for inhibiting RNA methylation
RI Glazer and MC Knode
The mechanism of action of the adenosine analog, neplanocin A (NPC), was
investigated in human colon carcinoma cell line HT-29. Cell viability was
reduced to 38 and 17% of control by 24-h exposure to 10(- 5) and 10(-4) M
NPC, respectively. Cytocidal activity was not affected by inhibition of
adenosine deaminase with 2'-deoxycoformycin. Concomitant with decreased
cell viability was the reduced incorporation of [14C]dThd and [3H]Leu, and
to a lesser extent [3H]Urd, into acid- precipitable material. Labeling of
rRNA and tRNA during drug treatment for 24 h with [methyl-3H]Met and
[14C]Urd revealed that NPC primarily inhibited RNA methylation, and to a
lesser extent, RNA synthesis. RNase T2 digests of total RNA indicated that
base and 2'-O-methylation were inhibited to approximately the same degree.
Metabolites of NPC were measured by reverse-phase high-performance liquid
chromatography and it was found that the major drug metabolite was the drug
analog of S- adenosylmethionine with little formation of the respective, S-
adenosylhomocysteine metabolite. NPC was utilized to a very small degree
for RNA synthesis where only 2 and 30 pmol of NPC/A260 were incorporated
into rRNA and tRNA after 24-h exposure to 10(-5) and 10(- 4) M NPC,
respectively. These results indicate that NPC is metabolized to a
metabolite of S-adenosylmethionine which is a poor methyl donor for RNA
methyltransferases, and that the accompanying decrease in RNA methylation
and protein synthesis appears to be related to its cytocidal activity.
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