Synthesis and evaluation of methylene blue oligonucleotide conjugates for DNA interstrand cross-linking

• Furan-modified oligonucleotides were prepared.
• Different photosensitizers were tested for singlet oxygen induced DNA crosslinking.
• DNA interstrand crosslinking is inefficient at low concentrations.
• Methylene blue was attached to complementary oligonucleotides.
• Using MB conjugates crosslinking could be achieved at low concentrations.

Efficient DNA interstrand cross-linking can be achieved with furan containing oligonucleotide probes upon activation by singlet oxygen (1O2). Previously, we have described how this can be achieved by irradiation of these furan probes with visible light in the presence of photosensitizers. Now, in an effort to explore cross-linking under conditions that are representative for experiments in cellular context, the furan mediated oligonucleotide cross-linking was investigated at low oligonucleotide concentrations, ensuring a sufficiently high local concentration of singlet oxygen by attaching the sensitizing methylene blue moiety to the oligonucleotide complementary to the furan modified strand. Four methylene blue-oligonucleotide conjugates were synthesized, each with a different positioning of methylene blue with respect to the furan unit present on the complementary strand. The conjugates were evaluated for singlet oxygen generation and subsequent cross-linking ability. It was observed that not only the distance of the 1O2 source to the furan unit, but also the specific interaction of methylene blue moiety with the duplex, which is position dependent, influences cross-linking yields.

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