Development of a method to isolate and analyze low abundance/low molecular weight proteins/peptides in the serum of breast cancer patients
Authors: T. Maity, D. Kirchner, S. Russell, V. S. K. Kolli, H. Brzeski, C. D. Shriver, Michael Liebman
Journal: Cancer Research (2005)

4821  Oxaliplatin (OX) is a second-generation platinum drug recently approved by the FDA for the treatment of colorectal cancer. OX differs structurally from cisplatin (CP), a widely used anticancer drug, by addition of a cyclohexane ring to the amines of CP to form a diaminocyclohexane (DACH) ring. Both CP and OX are known to bind to adjacent GG sequences in DNA. However, the molecular basis for the observed differences in their anticancer activity is not known. It is likely that the structure and/or conformational flexibility of CP-GG adducts differs from OX-GG adducts because of the presence of the bulky OX-DACH ligand in the major groove. To examine whether structural differences exist between OX and CP-DNA adducts, NMR solution structures of OX-adducted, CP-adducted and undamaged dodecamers were determined in the same sequence context, d(CCTCAGGCCTCC)•d(GGAGGCCTGA-GG). The NMR studies of OX and CP dodecamer adducts reveal certain similarities between them. For example, based on the H1-imino NOE cross-peak volume, and water and temperature dependence of the H1-imino cross-peak volume, it can be concluded for both dodecamers, that the G6 H1 was more solvent accessible than that of the G7. When the CP-GG and OX-GG adducts were compared to the undamaged dodecamer, the G6 and G7 H8 signals of both adducts shifted downfield. When the OX-GG and CP-GG dodecamer adducts were compared to CP-GG adducts with different sequence contexts, the OX and CP dodecamer adducts showed smaller downfield chemical shifts for the G6 H8 signal and no upfield shift for the A5 H2` signal. These results suggest that the orientation and/or chemical environment at the 5`-AGG-3` steps is affected by both the platinum and the sequence context. However, certain structural differences are also observed between the OX and CP dodecamer adducts around the central four base pairs (AGGC•GCCT) surrounding the adduct site. For example, while the G6 H8 protons of the CP-GG and OX-GG adducts had similar chemical shifts (8.27, 8.34 ppm, respectively), the differences in the chemical shift for the G7 H8 protons were larger (8.24, 8.02 ppm, respectively). This result suggests a difference in the G6 and G7 base canting for the CP- and OX-GG adducts. The calculated structures of the CP and OX dodecamer adducts show differences in twist at the A5-G6 and G6-G7 base pair steps and slide, tilt, and roll at the G6-G7 base pair step. Two ns unrestrained, fully solvated MD simulations of the same 12-mer OX- and CP- GG adduct have been completed, and are in good agreement with the NMR data. The MD simulations suggest that the mobility may be less for OX-GG than for CP-GG. These conformational and dynamic differences may help explain the differential recognition of OX and CP adducts by a number of cellular proteins and the resulting differences in cytotoxicity and mutagenicity of CP and OX adducts. (Supported by NIH grant CA84480).