GRANT NUMBER DAMD 17 - 94 - J - 4315 TITLE : Multiple Genetic Alterations in Breast Cancer

In an attempt to study the involvement of tyrosine kinase pathway in estrogen receptor signaling pathway, we used emodin, a tyrosine kinase inhibitor, to treat MCF-7 cells, a breast cancer cell line which express high level of estrogen receptor protein. Here we reported that emodin inhibited estrogen-induced expression of bcl-2 protein in MCF-7 breast cancer cells. Unexpectedly, treatment of the cells with emodin rapidly depletescellular levels of estrogen receptor protein in a doseand time-dependent manner. The pulse chase experiment showed that the decrease was resulted from enhanced degradationof estrogen receptor protein, not the rate of synthesis. To examine the mechanism involvedin the emodin-enhanced degradation of estrogen receptor, inhibitors of the lysosomal (chloroquine), proteasome (PSI and MG115), and calpains pathways were used. We found that only PSI and MG115, which specifically inhibit the chymotrypsin-like activityof proteasome, blocked emodin induced depletion of estrogen receptor protein levels. Theresults suggest that the proteasome proteolytic pathway may be involved in the emodin-induced decreases in estrogen receptor protein levels. We then examined the effect ofemodin on the hsp90-estrogen receptor heteromeric complex formation. We found that there was a marked increase in the complex formation. The data demonstrate that emodinmay inhibit the dissociation of hsp90 from estrogen receptor, resulting in the degradation ofestrogen receptor. Overall these findings indicate that proteasome-mediated proteindegradation can modulate estrogen receptor protein level, hsp90 may involve thedegradation of the estrogen receptor, and the possible use of emodin in the therapeutic manipulation of this process.