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BRCA1 has been implicated in a number of cellular processes, including transcriptional regulation, DNA damage repair, cell cycle arrest, and apoptosis. We identified mitogen-activated protein kinase (MAPK) kinase kinase 3 (MEKK3), an upstream regulator of the c-Jun NH(2)-terminal kinase/stress-activated protein kinase and p38/MAPK pathways, as a novel BRCA1-interacting protein in a yeast two-hybrid screen and confirmed the interaction by coimmunoprecipitation in mammalian cells. Deletion mapping demonstrated that amino acids 1611-1863 are required to mediate the interaction with MEKK3 in yeast. BRCA1 disease-associated mutations abrogated the interaction in yeast, and BRCA1 failed to interact with MEKK3 in BRCA1 mutant HCC1937 breast cancer cells. We demonstrate that small interfering RNA-based inhibition of endogenous BRCA1 reduces MEKK3 kinase activity and conversely that inducible expression of BRCA1 activates MEKK3 and p38/MAPK. Finally, we demonstrate using complementary approaches that BRCA1 is required for paclitaxel-induced activation of MEKK3. These data indicate that BRCA1 is a key regulator of the paclitaxel-induced stress response pathway and suggest that the ability of BRCA1 to associate with, and mediate the activation of, MEKK3 represents a potential mechanism through which this pathway is regulated.

Urokinase plasminogen activator (uPA) expression in breast cancer is associated with relapse and a reduction in disease-specific survival. Thus, efforts are under way to identify uPA inhibitors. By screening a chemical library of >1000 compounds, 17-allyaminogeldanamycin (17AAG) was identified as a potent inhibitor of uPA by the National Cancer Institute and is now in Phase I clinical trials. At this time, it remains unclear how 17AAG blocks uPA; one possibility is through disruption of the insulin-like growth factor I receptor (IGF-IR) pathway. This would be consistent with studies from our laboratory showing that activation of IGF-IR results in the induction of uPA protein. In the study described herein, we observed that IGF-IR and uPA were highly expressed in 87 and 55% of breast cancer by screening tumor tissue microarrays representing 930 cases. A significant proportion (52.1% = 354 of 680 cases, P < 0.0001) of the patients had tumors expressing both proteins. uPA alone (P = 0.033) or in combination with IGF-IR (P = 0.0104) was indicative of decreased disease-specific survival. Next, we demonstrated that treating MDA-MB-231 cells with increasing concentrations of 17AAG resulted in IGF-IR degradation (IC(50) = 1.0 micro M) and blocked signal transduction through the Akt and mitogen-activated protein kinase pathways. Finally, we found that 17AAG had a robust inhibitory effect on the production of uPA mRNAand protein in the presence of IGF-I. Thus, our study raises the possibility that 17AAG could prove to be an effective therapeutic agent for a large number of breast cancer patients by inhibiting the IGF-IR and ultimately uPA.