DESCRIPTION (provided by applicant): Of the platinum complexes currently utilized in cancer chemotherapy oxaliplatin (OX) appears to be less mutagenic than cisplatin (CP) or carboplatin. The mechanism(s) for these differences is not clear, but their mutagenicity is likely to be determined in part by DNA polymerase(s) that catalyze translesion synthesis past Pt-DNA adducts and/or damage-recognition proteins that bind to the adducts and prevent translesion synthesis. We postulate that there are differences in conformation and/or conformational flexibility between CP and OX adducts that affect the efficiency and fidelity of translesion synthesis. We have recently obtained the NMR solution structure for the OX-GG adduct in the mutagenic AGG sequence context, and it is significantly different from all previous solution structures of the CP-GG adduct. Thus, we plan to solve the NMR solution structure of the CP adduct and conduct NMR relaxation studies on OX and CP adducts in the AGG sequence context (aim 1). Our in vitro studies show that pol eta, pol beta, and pol mu are the only DNA polymerases with a significant capacity to bypass CP and OX adducts. Our data show that pol eta expression decreases the mutagenicity of CP, and previous studies have shown that pol beta overexpression increases the mutagenicity of CP. Pol beta is an excellent model for studying translesion synthesis because of the wealth of available structural and kinetic information. HMG-domain proteins such as SRY bind to CP and OX adducts with different affinities and can block translesion synthesis. Thus, we plan to extract NMR structural and dynamic information for OX and CP adducts in complex with pol beta and the damage-recognition protein SRY (aim 2); utilize molecular dynamics to characterize the impact of CP and OX adducts on DNA flexibility and to characterize the interaction of these adducts with pol beta (aim 3); and characterize the pre-steady state kinetics of pol beta with templates containing OX and CP adducts (aim 4). These data will also be used to verify and refine the molecular dynamic simulations performed in aim 3. These experiments will help define the mechanism(s) of Pt drug-induced mutagenesis and will result in models of Pt-DNA-polymerase interactions that will be useful in clarifying the mechanisms of translesion synthesis past bulky DNA adducts.
|Effective start/end date||3/1/08 → 2/28/10|
- National Cancer Institute: $279,826.00