Figure 2

Characterization of drug-DNA adducts. (a and b) Deconvoluted electrospray ionization mass spectrometry spectra of DNA (sgc8-5T-biotin) (a) and its drug-DNA adducts with doxorubicin (b). Compared with DNA, the peaks (drug-DNA adducts 1–6) shifted to larger molecular weights in drug-DNA adducts (b), corresponding to drug-DNA adducts with 1–6 copies of doxorubicin, respectively, conjugated on one DNA (Supplementary Table S2). (5 T: a spacer; Biotin: for streptavidin conjugation in following study) (c) UV–Vis absorption spectra of drug-DNA adducts and control (reaction solution without formaldehyde). (d) Fluorescence spectra of free Dox (2 μM), drug-DNA adducts (2-μM doxorubicin equivalents) and control. (e) High stability of drug-DNA adducts at 4 °C and conditional drug release at a physiological temperature (37 °C). (Solid lines: fit curves) (f) An agarose gel electrophoresis image indicating that drug-DNA adducts (2-μM DNA equivalent) were resistant to nuclease (0.05-U μl⁻¹ DNase I) degradation. Ethidium bromide was used for DNA staining. The weaker intensity in lane 1 compared with that in lanes 3 or 4 is likely because of the energy transfer from ethidium bromide to conjugated doxorubicin in drug-DNA adducts. (lane legends: 1, DNA; 2, DNA+DNase I; 3, drug-DNA adducts; 4, drug-DNA adducts+DNase I; and 5, DNA marker).