SEQUENCE-SELECTIVE GUANINE REACTIVITY BY DUOCARMYCIN-A

High selectivity for covalent reaction at adenine N-3 within duplex DN A is a distinguishing feature of the CC-1065 and duocarmycin classes o f natural products. Studies of the base and sequence selectivity exhib ited by duocarmycins and CC-1065-based alkylating agents have focused on characterization of the predominant covalent adenine adducts that a re formed. While information about minor DNA reaction products could p rovide valuable insights to our understanding the DNA recognition and reactivity properties of these agents, little characterization of such adducts by these agents has appeared in the literature. To broaden ou r structure-reactivity understanding of these DNA alkylating compounds , comparative investigations of the covalent sequence selectivity exhi bited by compounds containing altered cyclopropapyrroloindole (CPI) al kylating subunits such as duocarmycin A were undertaken using the DNA polymerase inhibition assay. We were surprised to identify with this a ssay a DNA sequence with an unusual propensity for covalent reaction w ith duocarmycin A at a guanine nucleotide. Using the heat strand break age assay with a duplex oligonucleotide containing this interesting se quence, we confirmed the site of alkylation to be the indicated guanin e in the sequence 5'-CGCGTTGGGAG-3'. The trimethoxyindole-CPI analog of duocarmycin A does not alkylate this guanine, suggesting that there are interesting features to the duplex recognition/reactivity exhibit ed by duocarmycin A. Herein we describe our identification of the firs t DNA sequence which covalently reacts with duocarmycin A at a guanine nucleotide in the absence of additional minor groove binding agents.