The Antileukemia Drug 2-Chloro-2′-deoxyadenosine: An Intrinsic Transcriptional Antagonist

Document Type

Article

Publication Title

Molecular Pharmacology

Abstract

The nucleoside analog 2-chloro-2'-deoxyadenosine (CldAdo; cladribine) is effective in the treatment of hairy cell leukemia and chronic lymphocytic leukemia. CldAdo is phosphorylated and incorporated into cellular DNA but is not an absolute chain terminator. We demonstrated by in vitro gel-shift assays that binding interactions of the human TATA box-binding protein (TBP) were disrupted on 2-chlorodeoxyadenosine monophosphate (CldAMP)-substituted TATA box consensus sequences. We hypothesized that human RNA polymerase II (pol II) transcriptional processes would therefore be affected by 2-chlorodeoxyadenosine triphosphate (CldATP) incorporation into a promoter TATA element. Double-stranded DNA templates containing the adenovirus major late promoter and coding sequences were enzymatically synthesized as control or with site-specific CldAMP residues, incubated with HeLa extract, and the synthesis of radiolabeled 44-base transcripts was assessed. With increasing amounts of HeLa extract, CldAMP substitution for dAMP within the TATA box decreased in vitro pol II transcription by approximately 35% compared with control substrates. Time-course studies showed that transcript production increased in a linear fashion on control substrates. In contrast, transcription on CldAMP-substituted TATA sequences reached a plateau after 20 min. Furthermore, CldAMP-substituted promoter sequences trapped or sequestered TBP, preventing its dissociation from DNA and subsequent binding to additional TATA elements to reinitiate transcription. CldAdo thus represents the first example of a nucleoside analog that acts as a transcriptional antagonist. CldATP incorporation into gene regulatory sequences may provide a novel strategy to modulate specific protein/DNA interactions.

DOI

10.1124/mol.65.1.227

Publication Date

1-2004

ISSN

1521-0111

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