(Application Abstract) P-glycoprotein (Pgp) was first identified by virtue of its overexpression in multidrug-resistant cells, where it mediates the efflux of a large number of chemotherapeutic agents. Moreover, recent evidence suggests that Pgp may also play a more general anti-apoptotic role in tumor cells, since cells overexpressing Pgp also exhibit resistance to a number of other caspase-mediated apoptotic inducers, including serum starvation, UV irradiation, Fas ligand and TNF. Interestingly, many of these agents also act as inducers of Pgp transcription, suggesting that activation of Pgp may be part of a general """"""""stress response"""""""" of tumor cells, and play a role in cellular growth and death decisions in response to toxic stimuli. Our laboratory has had a long-standing interest in the mechanisms regulating the transcription of the human Pgp gene, MDR1. Our recent observation that MDR1 gene expression can be rapidly (within minutes) activated within patient tumors exposed to the genotoxic chemotherapeutic agent doxorubicin has prompted us to investigate the mechanism underlying transcriptional induction of MDR1 by a variety of """"""""stress"""""""" agents. Surprisingly, we have found that signals from seemingly disparate agents (including chromatin modifiers, differentiation agents, chemotherapeutics and UV irradiation) converge on a small region of the MDR1 promoter, and have found that this region interacts with a multisubunit complex, which we refer to as the """"""""MDR1 enhancesome,"""""""" that includes the transcription factors NF-Y, Sp1 and PCAF. Furthermore, we have identified a novel chemotherapeutic agent, ET-743, which inhibits activation of MDR1 through the enhancesome, without significantly affecting constitutive MDR1 transcription. We now propose to continue our investigations of the """"""""MDR1 enhancesome"""""""" and ET-743: 1) To identify and characterize additional components of the """"""""MDR1 enhancesome""""""""; 2) To investigate the effect of transcriptional inducers on chromatin remodeling at the MDR1 promoter; 3) To pursue our finding that multiple Sp1 family members may be components of the MDR1 enhancesome and have divergent effects on MDR1 transcription and 4) To continue to investigate the mechanism by which ET-743 inhibits activation of MDR1 transcription, with the long-term goal of identifying the specific transcriptional target of this novel chemotherapeutic agent.
