Akt and ERK activation in relation to acquired hormone resistance and metastatic potential in a murine breast cancer model

WARGON VICTORIA; GOROSTIAGA MARIA; NOVARO VIRGINIA; LANARI CLAUDIA

SAN DIEGO, USA

Congreso; AACR; 2008

AACR

The C4-HI tumor is a murine hormone-independent (HI) mammary carcinoma that expresses estrogen receptors (ER) and both progesterone receptor (PR) isoforms (PR A and PR B); it is maintained by syngeneic transplantations and regresses after antiprogestin treatment. We have developed a tumor variant which acquired resistance to the antiprogestin RU 486, C4-HIR, that  expresses lower PR A levels than C4-HI (p<0.05), and is more metastatic than the parental tumor. We have also demonstrated that acquired hormone resistance is a reversible phenomenon and we generated a new variant, C4-HIRrev that re-expressed PR A. Curiously, the metastatic phenotype acquired by C4-HIR was not reverted. On the other hand, in previous studies, we have demonstrated that carcinoma associated fibroblasts (CAF) from HI and hormone dependent tumors were different. The aim of this study was a) to investigate the activation of Akt (p-Akt) and Erk (p-Erk) in C4-HI, C4-HIR and C4-HIRrev tumors in vivo and b) to evaluate the role of CAF in the hormone resistant phenotype. Tumor extracts from the three tumor variants were used to analyze Akt and ERK activation by Western-Blot. We observed an increased activation in Akt (pAkt/Akt) in C4-HI as compared with C4-HIR and C4-HIRrev (p<0,05). Immunohistochemical studies using the pSer 473 and pThr 308 antibodies, confirmed higher nuclear staining in C4-HI samples. The opposite occurred with p-Erk/Erk expression, which was higher in C4-HIR and C4-HIRrev as compared with C4-HI (p<0,05) suggesting that the pattern of pAkt and pErk activation is related to the metastatic behavior rather than to tumor hormone responsiveness. In addition, primary cell cultures of C4-HI and C4-HIR were performed and CAF were separated from epithelial tumor cells (EPI). The responsiveness of the purified EPI (C4-HI or C4-HIR) and of the co-cultures (EPI + CAF) to RU 486 was evaluated. Surprisingly, EPI from both tumors were similarly inhibited with antiprogestins and this inhibition was even stronger in the presence of CAF from C4-HIR or C4-HI. The lack of differences in RU 486 responsiveness between both tumors in vitro, led us to evaluate PR expression in cell cultures. No differences in PR A expression were detected between C4-HI and C4-HIR growing in plastic, confirming that the ability to respond to antiprogestins is dependent on PR A expression. In summary these results suggest that PR A expression may be regulated by epigenetic mechanisms reverted by plastic attachment, and that in this model low p-Akt and high p-Erk are related with a high metastatic potential and not with their hormone responsiveness.