Molecular Assembly of Triplex of Duplexes from HomothyminylHomocytosinyl Cγ(S/R)-Bimodal Peptide Nucleic Acids with dA8/dG6 and the Cell Permeability of Bimodal Peptide Nucleic Acids

Peptide nucleic acids (PNAs) are analogues of DNA with a neutral acyclic polyamide backbone containing nucleobases attached through a t-amide link on repeating units of aminoethylglycine (aeg). They bind to complementary DNA or RNA in a sequencespecific manner to form duplexes with higher stablity than DNA:DNA and DNA:RNA hybrids. We have recently explored a new type of PNA termed bimodal PNA (bm-PNA) designed with two nucleobases per aeg repeating unit of PNA oligomer and attached at Ca or C gamma of each aeg unit through a spacer sidechain. We demonstrated that C gamma-bimodal PNA oligomers with mixed nucleobase sequences bind concurrently two different complementary DNAs, forming double duplexes, one from each t-amide and C gamma face, sharing a common PNA backbone. In such bm-PNA:DNA ternary complexes, the two duplexes show higher thermal stability than individual duplexes. Herein, we show that C gamma(S/R)-bimodal PNAs with homothymines (T-8) on a t-amide face and homocytosine (C-6) on a C gamma-face form a conjoined pentameric complex consisting of a triplex (bm-PNA-T-8)(2):dA(8) and two duplexes of bm-PNA-C-6:dG(6). The pentameric complex [dG(6):C gamma(S/R)-bm-PNA:dA(8):C gamma(S/R)-bm-PNA:dG(6)] exhibits higher thermal stability than the individual triplex and duplex, with C gamma(S)-bm-PNA complexes being more stable than C gamma(R)-bm-PNA complexes. The conjoined duplexes of C gamma-bimodal PNAs can be used to generate novel higher-order assemblies with DNA and RNA. The C gamma(S/R)-bimodal PNAs are shown to enter MCF7 and NIH 3T3 cells and exhibit low toxicity to cells.