Linkers designed to intercalate the double helix greatly facilitate DNA alkylation by triplex-forming oligonucleotides carrying a cyclopropapyrroloindole reactive moiety

Tripler-forming oligonucleotides (TFOs) bind sequence-specifically in the m ajor groove of double-stranded DNA. Cyclopropapyrroloindole (CPI), the elec trophilic moiety that comprises the reactive subunit of the antibiotic CC-1 065, gives hybridization-triggered alkylation at the N-3 position of adenin es when bound in the minor groove of double-stranded DNA. In order to attai n TFO-directed targeting of CPI, we designed and tested linkers to 'thread' DNA from the major groove-bound TFO to the minor groove binding site of CP I, Placement of an aromatic ring in the linker significantly enhanced the s ite-directed reaction, possibly due to a 'threading' mechanism where the ar omatic ring is intercalated. All of the linkers containing aromatic rings p rovided efficient alkylation of the duplex target. The linker containing an acridine ring system, the strongest intercalator in the series, gave a sma ll but clearly detectable amount of non-TFO-specific alkylation, An equival ent-length linker without an aromatic ring was very inefficient in DNA targ et alkylation.