Steroid-induced androgen receptor-oestradiol receptor beta-Src complex triggers prostate cancer cell proliferation

Abstract

Treatment of human prostate carcinoma-derived LNCaP cells with androgen or oestradiol triggers simultaneous association of androgen receptor and oestradiol receptor b with Src, activates the Src/Raf-1/ Erk-2 pathway and stimulates cell proliferation. Surprisingly, either androgen or oestradiol action on each of these steps is inhibited by both anti-androgens and anti-oestrogens. Similar ®ndings for oestradiol receptor a were observed in MCF-7 or T47D cells stimulated by either oestradiol or androgens. Microinjection of LNCaP, MCF-7 and T47D cells with SrcK ± abolishes steroid-stimulated S-phase entry. Data from transfected Cos cells con®rm and extend the ®ndings from these cells. Hormone-stimulated Src interaction with the androgen receptor and oestradiol receptor a or b is detected using glutathione S-transferase fusion constructs. Src SH2 interacts with phosphotyrosine 537 of oestradiol receptor a and the Src SH3 domain with a proline-rich stretch of the androgen receptor. The role of this phosphotyrosine is stressed by its requirement for association of oestradiol receptor a with Src and consequent activation of Src in intact Cos cells.

Inhibition of the SH3 domain-mediated binding of Src to the androgen receptor and its effect on tumor growth

“…In the presence of the synthetic androgen R1881, a strong association of wt AR with Src was detected, whereas a much weaker association, similar to that found in the absence of hormone, was observed in cells expressing the AR mutants (Figure 1b, lower panel). As expected (Migliaccio et al, 2000), Src immunoprecipitated with the wt AR from the lysate of Cos cells treated with R1881 for 3 min actively phosphorylated acidified enolase. In contrast, no increase of Src activity over the basal level was observed when the AR constructs with a deletion of either the 372-385 residues (ARÀD1) or the 372-378 sequence (ARÀD2) were overexpressed in the same cells (Figure 1c).…”

“…However, this approach is limited by the mixed, antagonist and agonist action of these molecules, their side effects, and the appearance of hormone resistance. Estradiol, progestins and androgens induce G1 to S transition of cells derived from human mammary and prostate cancers through activation of signal transducing pathways (Castoria et al, 1999;Migliaccio et al, 2000). These findings open potential new approaches to the therapy of hormonedependent cancers.…”

“…Deletion of the 371-422 sequence of the hAR abolishes the in vitro association of the receptor with the Src SH3 domain (Migliaccio et al, 2000). To better define the AR-interacting sequence, different human AR-deletion mutants were prepared.…”

“…On the basis of our previous findings (Migliaccio et al, 2000) and establishment of new functional assays for AR deletion mutants, in this report, we define the amino-acid sequence responsible for the human AR (hAR) interaction with Src. A synthetic 10 amino-acid peptide that mimics this sequence prevents the AR/ER complex from associating with Src.…”

“…In human prostate and mammary cancer-derived cells, treatment with androgen or estrogen rapidly induces association of Src with AR and ER, leading to Src activation and stimulation of DNA synthesis (Migliaccio et al, 2000). AR association with Src has also been observed in androgen-stimulated immortalized fibroblasts (Castoria et al, 2003), raising the possibility that, in response to androgens, stromal cells facilitate the malignant outgrowth and the metastatic spread of prostate epithelial cells.…”