Quinone Methides Tethered to Naphthalene Diimides as Selective G-Quadruplex Alkylating Agents

TLDR

The study develops G‑quadruplex ligand/alkylating hybrids that tether naphthalene diimides to Mannich‑base quaternary ammonium salts, generating quinone‑methide precursors activatable by mild heating. The bis‑substituted naphthalene diimides were synthesized and shown to act as activatable bis‑alkylating agents; the quinone‑methide intermediate was trapped via a hetero Diels–Alder reaction, and DNA binding and alkylation were probed by gel electrophoresis, circular dichroism, and enzymatic assays. The hybrids alkylated G‑quadruplex DNA preferentially over other conformations, and reversible recognition was found to be a prerequisite for alkylation, thereby reinforcing G‑quadruplex structural rearrangement.

Abstract

We have developed novel G-quadruplex (G-4) ligand/alkylating hybrid structures, tethering the naphthalene diimide moiety to quaternary ammonium salts of Mannich bases, as quinone-methide precursors, activatable by mild thermal digestion (40 °C). The bis-substituted naphthalene diimides were efficiently synthesized, and their reactivity as activatable bis-alkylating agents was investigated in the presence of thiols and amines in aqueous buffered solutions. The electrophilic intermediate, quinone-methide, involved in the alkylation process was trapped, in the presence of ethyl vinyl ether, in a hetero Diels−Alder [4 + 2] cycloaddition reaction, yielding a substituted 2-ethoxychroman. The DNA recognition and alkylation properties of these new derivatives were investigated by gel electrophoresis, circular dichroism, and enzymatic assays. The alkylation process occurred preferentially on the G-4 structure in comparison to other DNA conformations. By dissecting reversible recognition and alkylation events, we found that the reversible process is a prerequisite to DNA alkylation, which in turn reinforces the G-quadruplex structural rearrangement.