Androgen regulation of TNFR family-induced apoptosis

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The progression from androgen dependent to androgen independent disease is a critical area of investigation since effective regimens to treat androgen independent disease are not available. Our long-term goal is to understand the mechanisms of resistance to apoptosis in prostate cancer because apoptosis is an attractive therapeutic target in this setting. We have recently identified cell death pathways that are regulated by androgen/AR and that could have important implications. It has been reported that LNCaP displays a bell-shaped growth curve in response to increasing doses of DHT. Low doses DHT (in the pM range) result in an increase in cell proliferation whereas higher doses (in the nM-mM range) result in growth arrest. We have found that this bell-shaped growth curve was dramatically changed after androgen deprivation: culturing of LNCaP for 10 days in SFC resulted in an increase in cell proliferation after treatment with high doses of DHT. We concluded that after androgen deprivation the AR responds differently to androgen treatment but the mechanism of this different response is unknown. In LNCaP TNF, as well as TRAIL, and anti-Fas agonistic Ab-induced apoptosis (in the presence of wortmannin) were inhibited by DHT, although to varying degrees. Caspase activity, as measured by a fluorogenic assay or western blot, was inhibited but was dependent on the specific caspase, growth in FCS or SFC, and the concentration of DHT. TRAIL R1 and R2 expression decreased in steroid free media (SFC) and could be restored with DHT. SFC increased the level of activated (phospho) Akt. Treatment with the PI3K inhibitor wortmannin indicated that the inhibitory effect is downstream from the DISC and dependent on the PI3K-Akt pathway. In FCS mRNA (northern blot) and protein (western blot) levels of p53 decreased with DHT and there was significant effect on p53 when grown in SFC on protein levels but not at the level of mRNA. Caspase activity was decreased in LNCaP-sip53 and DHT further decreased this activity. PC3 transfected with wt AR was relatively resistant to ligand induced cell death. DHT regulated caspase 2 levels and chromatin immunoprecipitation analysis indicated that there is direct interaction between AR and caspase-2 gene. Taken together, our data strongly suggest a role for the androgen milieu in regulating TNFR ligand-induced apoptosis that has important clinical implications in patients who have previously been treated with ADT.