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Roles of Chk1 Kinase in Breast Cancer Etiology and Therapy
Background: S phase checkpoints insure the fidelity and timing of DNA replication in proliferating cells. Thus, attacking these surveillance pathways provides us new routes for killing solid tumors, including breast cancers. This strategy appears to be particularly effective against hypoxic solid tumors, due in part to the fact that tumor hypoxia imposes particular stress on the process of DNA replication. We have shown that S-phase specific genotoxins induce the ubiquitin-dependent degradation of the checkpoint kinase Chk1 in breast cancer cells. Downregulation of Chk1 is centrally involved in the cytotoxic mechanism of these agents. Breast cancer cells that fail to degrade Chk1 exhibit marked resistance to these genotoxins. Thus, a more comprehensive understanding of the Chk1 ubiquitination pathway will yield new insights into breast cancer etiology and therapy. Moreover, because hypoxic stress is a major environmental source of S-phase genotoxicity in cancers, the ATR-Chk1-dependent checkpoint pathway is predicted to play major roles in the maintenance of DNA replication fidelity and cell viability in hypoxic tumor cells. Objective/Hypothesis: This project is designed to understand the roles of Chk1 degradation in breast cancer etiology and therapy. As an integrated objective, we will explore the hypothesis that defects in Chk1 degradation pathway contribute to resistance to anti-cancer agents in breast cancer cells. Specific Aims: 1. To further characterize the molecular components of the Chk1 degradation pathway. 2. To investigate the hypotheses that defects in Chk1 degradation contribute to resistance to both anti-cancer drug and hypoxia, and that inhibition of Chk1 represents a novel therapeutic strategy for killing of hypoxic breast cancer cells. Study Design: For Aim 1, a proteomic approach will be used to identify Chk1-binding components of the E3 ligase complexes that target Chk1 for degradation. We will further characterize functions of these proteins in breast cancer cells. For Aim 2, we will explore the effect of hypoxia on the Chk1 destruction pathway, will examine the contributions of defects in this pathway to both hypoxia and drug resistance in breast cancer cells. Potential outcomes and Benefits of the Research: Characterization of the Chk1 destruction machinery will provide novel insights into the causes of genetic instability and anticancer drug resistance in breast cancer cells. Furthermore, this line of research has direct implications for the development of novel therapeutic agents targeted against breast cancer and other solid tumors.
Roles of Chk1 Kinase in Breast Cancer Etiology and Therapy.
Most solid tumors, including breast cancers, grow in a highly stressful micro-environment characterized by a severely limited blood supply and abnormally low levels of oxygen (termed hypoxia). Severe oxygen depletion halts the copying of the genome (termed DNA replication) in proliferating cells by activating a protective mechanism termed the S-phase checkpoint. Moreover, this checkpoint also allows the tumor to actively resume DNA replication when environmental conditions, including the oxygen supply, are more favorable to support this energy-intensive process. Our underlying hypothesis is that breast tumor cells rely heavily on the S phase checkpoint to maintain both DNA replication and cell proliferation in human tissues. We have shown that certain anticancer drugs trigger the destruction of the S phase checkpoint kinase protein Chk1 in breast cancer cells, and that this effect is critical for the cancer cell-killing activity of some of our most clinically useful anticancer agents. Of equal importance, we have shown that the failure of these cells to degrade Chk1 leads to drug resistance, which translates to tumor relapse and treatment failure in the clinical setting. The hypoxic regions of solid tumors are considered to be a breeding ground for increasingly malignant, drug-resistant cancer cells ¨C the same cells that give rise to tumor re-appearance as well as metastatic disease. Based on our work to date, we hypothesize that highly malignant breast cancer cells may have defects in the Chk1 degradation machinery, which not only allows cell proliferation under hypoxic conditions, but also confers resistance to some important anticancer agents. The goals of this project are to further define the destruction machinery for Chk1 in breast cancer cells and to test the hypothesis that breast cancer cells may have defects in this Chk1 destruction machinery. We will also examine whether manipulation of the Chk1 destruction pathway represents a feasible therapeutic approach that will allow oncologists to effectively kill the most life-threatening cells in the breast tumor mass ¨C those cells that are surviving under conditions of severe oxygen and nutrient depletion.