Comparative effects of 3'-azido-3'-deoxythymidine and its metabolite 3'-amino-3'-deoxythymidine on hemoglobin synthesis in K-562 human leukemia cells.
We previously demonstrated that 3'-azido-3'-deoxythymidine (AZT) inhibits hemoglobin (Hb) synthesis and globin gene transcription in butyric acid-induced K-562 leukemia cells, suggesting that these effects may play a role in the AZT-induced anemia observed in patients [Mol. Pharmacol. 38:797-804 (1990)]. The recent discovery by our group of a novel metabolite of AZT. 3'-amino-3'-deoxythymidine (AMT), which exhibits a high degree of toxicity toward human hemopoietic cells [Mol. Pharmacol. 39:258-266 (1991); Antimicrob. Agents Chemother. 35:801-807 (1991)], has led us to explore potential effects of this AZT metabolite on Hb production, globin mRNA expression, and heme synthesis in butyric acid-induced K-562 human erythroleukemia cells. AMT inhibited Hb synthesis by approximately 21%, as measured by benzidine staining, at concentrations as low as 25 microM, with slightly increased inhibition at higher AMT concentrations. The inhibition of Hb production by AMT was substantially lower, compared with that of AZT. AMT inhibited globin mRNA steady state levels in a dose-dependent manner to a similar extent as did the parent drug, with approximately 50% inhibition by each compound at a concentration of 100 microM. Nuclear run-on transcription assays demonstrated that inhibition by AMT of globin mRNA synthesis was associated with a decreased rate of globin-specific gene transcription. Globin mRNA stability was not affected by either 100 microM AZT or AMT, as measured after blockage of transcription with actinomycin D. To gain insight into potential mechanism(s) responsible for the different quantitative effects of AZT and AMT on Hb synthesis, the effect of each compound on induction of heme synthesis in K-562 cells was determined. Although heme induction was not affected by AMT, a significant inhibition approximating 20% was observed in the presence of 100 microM AZT. In addition, AZT down-regulated mRNA steady state levels under conditions where heme synthesis was inhibited by succinylacetone. These data suggest that inhibition by AZT of globin gene expression is a direct effect and is not secondary to inhibition of heme synthesis. This study emphasizes the role of AMT in the pharmacodynamic properties of AZT, in relation to its toxicity, and suggest that both AMT and AZT may be involved in the inhibition of erythroid differentiation observed in vivo, through changes in gene expression.
Weidner
D A
DA
Department of Pharmacology, University of Alabama, Birmingham 35294.
Bridges
E G
EG
Cretton
E M
EM
Sommadossi
J P
JP
D002087:Butyrates; D020148:Butyric Acid; D015224:Dideoxynucleosides / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D005914:Globins / Q000096:biosynthesis / Q000235:genetics; D006418:Heme / Q000378:metabolism; D006454:Hemoglobins / Q000096:biosynthesis*; D006801:Humans; D007942:Leukemia, Experimental / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects; D014407:Tumor Cells, Cultured; D015215:Zidovudine / Q000494:pharmacology*
D002087:Butyrates; D015224:Dideoxynucleosides; D006454:Hemoglobins; D012333:RNA, Messenger; D020148:Butyric Acid; D006418:Heme; D015215:Zidovudine; C019252:3'-aminothymidine; D005914:Globins