DNA crosslinks, including interstrand crosslinks (ICLs) and DNA-protein crosslinks (DPCs) are forms of DNA damage that arise continuously in DNA from endogenous and natural sources. They must be removed in order to allow accurate genome duplication and gene expression. Many chemotherapeutic agents induce ICLs including nitrogen mustards and derivatives (melphalan, chlorambucil), psoralens, mitomycin C, platinum- based compounds such as cisplatin, and nitrosoureas such as BCNU. Identification and future development of biomarkers associated with ICL repair proficiency will facilitate precision medical treatment for individual patients. Despite the importance of crosslinks in cancer etiology and treatment, mechanisms of DNA crosslink repair are known only in outline, and many steps are simply assumed. An integrated, programmatic approach involving four Research Projects and two service Cores will conduct experimental investigations under the auspices of the Program and contribute, both individually and synergistically, to this theme. Each Overall Goal/Specific Aim is independently engaged by each project using distinct and complementary approaches. The Program Project is organized around three aims, representing goals of the project:
Aim 1 : Determine how different pathways of crosslink repair are used or coordinated. The program will investigate major gaps in knowledge about different crosslink repair pathways, including how nucleases or complexes/components are recruited and used in different situations, variations of crosslink repair during the cell cycle, action of crosslink repair at different configurations of model stalled replication forks, whether ICL repair differs for different lesions, and the mechanism of error-free and mutagenic crosslink repair.
Aim 2 : Understand functions of newly discovered and little-studied crosslink repair components. One block to progress in understanding repair of ICLs is that there are a group of components that are known to be involved in crosslink repair, but where mechanistic roles are unassigned or known only superficially. In this program the investigators will cooperate to determine their functions. Major unknown components include SLX4IP (which associates with SLX4); the HMGB1, HMGB2 and HMGB3 DNA binding proteins; the MSH2-MSH3 complex (which binds DNA distortions); and UHRF1 (postulated as an alternative scaffold for delivery of nucleases).
Aim 3 : Investigate the mechanisms of repair of DNA-protein crosslinks. There are likely to be several routes of DPC repair and tolerance. Experiments will be undertaken to determine different modes of processing. Major gaps in knowledge include the involvement of Fanconi anemia (FA) pathway proteins in repair and how features of FA might be explained by defects in DPC repair. We will investigate involvement of replicative bypass (translesion DNA synthesis) in tolerance of DNA-peptide crosslinks (modeling proteolytically processed DPCs). We will also explore pathways that prevent or induce mutagenesis by DPCs.
Drugs such as cisplatin analogs, psoralens, and nitrogen mustard derivatives cause DNA crosslinks, which are effective in eliminating tumor cells. This Program Project Proposal describes a team with expertise in biochemistry, cell biology, and genetics, supported by two Core facilities, to clarifying key steps in DNA crosslink repair. Successful completion of this Program Project will define molecular pathways of DNA crosslink repair and identify novel targets that can be explored to improve chemotherapy.
