Oligonucleotides incorporating a central C‐nucleoside with either a rigid or flexible benzaldoxime base moiety have been synthesized, and the hybridization properties of their metallacyclic derivatives have been studied by UV melting experiments. In all cases, the metallated duplexes were less stable than their unmetallated counterparts, and the metallacyclic nucleobases did not show a clear preference for any of the canonical nucleobases as a base‐pairing partner. With palladated oligonucleotides, increased flexibility translated to less severe destabilization, whereas the opposite was true for the mercurated oligonucleotides; this reflects the greater difficulties in accommodating a rigid Pd II ‐mediated base pair than a rigid Hg II ‐mediated base pair within the base stack of a double helix.

Covalent metalation of the base moieties affords a new class of modified oligonucleotides. These organometallic oligonucleotides share many properties, notably increased hybridization affinity conferred by metal-mediated base pairing, with oligonucleotides incorporating coordinative transition-metal complexes. They are, however, set apart by their ability to retain the transition-metal ion even at extreme dilution. Such stability towards dissociation would be desirable in DNA nanotechnology and necessary in therapeutic applications. Herein we describe our efforts towards preparation and characterization of covalently mercurated and palladated oligonucleotides, highlighting in particular our recent contribution on the synthesis and potential applications of oligonucleotides incorporating dimercurated artificial nucleobases.1 Introduction2 Synthesis of Covalently Mercurated and Palladated Oligonucleotides3 Hybridization Properties of Covalently Mercurated and Palladated Oligonucleotides4 Outlook

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Short oligonucleotides with cyclopalladated benzylamine moieties at their 5′-termini have been prepared to test the possibility of conferring palladacyclic anticancer agents sequence-selectivity by conjugation with a guiding oligonucleotide. Hybridization of these oligonucleotides with natural counterparts was studied by UV and CD (circular dichroism) melting experiments in the absence and presence of a competing ligand (2-mercaptoethanol). Cyclopalladated benzylamine proved to be strongly stabilizing relative to unmetalated benzylamine and modestly stabilizing relative to an extra A•T base pair. The stabilization was largely abolished in the presence of 2-mercaptoethanol, suggesting direct coordination of Pd(II) to a nucleobase of the complementary strand. In all cases, fidelity of Watson-Crick base pairing between the two strands was retained. Hybridization of the cyclopalladated oligonucleotides was characterized by relatively large negative enthalpy and entropy, consistent with stabilizing Pd(II) coordination partially offset by the entropic penalty of imposing conformational constraints on the flexible diethylene glycol linker between the oligonucleotide and the palladacyclic moiety.

2’-O-Methylribo phosphorothioate oligonucleotides incorporating cyclopalladated benzylamine conjugate groups at their 5’-termini have been prepared and their ability to hybridize with a designated target sequence was assessed by conventional UV melting experiments. The oligonucleotides were further examined in splice-switching experiments in human cervical cancer (HeLa Luc/705), human liver (HuH7_705), and human osteosarcoma (U-2 OS_705) reporter cell lines. Melting temperatures of duplexes formed by the modified oligonucleotides were approximately 5 °C lower than melting temperatures of the respective unmodified duplexes. The cyclopalladated oligonucleotides functioned as splice-correcting agents in the HeLa Luc/705 cell line somewhat more efficiently than their unmodified counterparts. Furthermore, the introduction of this chemical modification did not induce toxicity in cells. These results demonstrate the feasibility of using covalently metalated oligonucleotides as therapeutic agents.