Genome-Wide Analysis of Aromatase Inhibitor-Resistant, Tamoxifen-Resistant, and Long-Term Estrogen-Deprived Cells Reveals a Role for Estrogen Receptor

Abstract

Acquired resistance to either tamoxifen or aromatase inhibitors (AI) develops after prolonged treatment in a majority of hormone-responsive breast cancers. In an attempt to further elucidate mechanisms of acquired resistance to AIs, MCF-7aro cells resistant to letrozole (T+LET R), anastrozole (T+ANA R), and exemestane (T+EXE R), as well as long-term estrogen deprived (LTEDaro) and tamoxifen-resistant (T+TAM R) lines were generated. This is the first complete panel of endocrine therapy-resistant cell lines, which were generated as multiple independent biological replicates for unbiased genome-wide analysis using affymetrix microarrays. Although similarities are apparent, microarray results clearly show gene signatures unique to AI-resistance were inherently different from LTEDaro and T+TAM R gene expression profiles. Based on hierarchical clustering, unique estrogen-responsive gene signatures vary depending on cell line, with some genes upregulated in all lines versus other genes up-regulated only in the AI-resistant lines. Characterization of these resistant lines showed that LTEDaro, T+LET R, and T+ANA R cells contained a constitutively active estrogen receptor (ER)A that does not require estrogen for activation. This ligand-independent activation of ER was not observed in the parental cells, as well as T+EXE R and T+TAM R cells. Further characterization of these resistant lines was performed using cell cycle analysis, immunofluorescence experiments to visualize ER subcellular localization, as well as cross-resistance studies to determine second-line inhibitor response. Using this well-defined model system, our studies provide important information regarding differences in resistance mechanisms to AIs, TAM, and LTEDaro, which are critical in overcoming resistance when treating hormone-responsive breast cancers. [Cancer Res 2008;68(12):4910-8]

Biological reprogramming in acquired resistance to endocrine therapy of breast cancer

“…As supported by the molecular data presented above, acquired resistance may be mediated by constitutive activation of ERa and growth factor signaling pathway cross-talk (Martin et al, 2003;Jelovac et al, 2005;Sabnis et al, 2005;Santen et al, 2005;Masri et al, 2008). Thus, the identities of genes differentially expressed during MCF7-LTED adaptation may support the important role of the non-genomic function of ERa.…”

“…Current literature supports the hypothesis that acquired resistance is mainly mediated by molecular events that-particularly in the case of resistance to AIs-lead to constitutive activation of ERa and growth factor signaling pathway cross-talk (Clarke et al, 2003;Martin et al, 2003;Yue et al, 2003;Jelovac et al, 2005;Normanno et al, 2005;Sabnis et al, 2005;Santen et al, 2005;Masri et al, 2008). On the basis of these studies, several clinical trials have attempted to overcome resistance through combination with growth factor signaling inhibitors Massarweh and Schiff, 2006).…”

SGK3 sustains ERα signaling and drives acquired aromatase inhibitor resistance through maintaining endoplasmic reticulum homeostasis

“…LTED cells do not respond to any AI, whereas our AI-resistant cells acquire resistance to one AI and will respond to another AI (16,32), which resembles a true clinical situation (33). Moreover, LTED cells are less responsive to fulvestrant compared with our AI-resistant cells (16), suggesting LTED cells are less dependent on ERα signaling than our resistant cell lines. Therefore, LTED cells and our acquired AI-resistant cells may represent different stages of AI resistance.…”

“…aromatase-overexpressing ERα + breast cancer cell line MCF7aro to testosterone (T) plus exemestane, anastrozole, and letrozole, respectively (16). Resistance to these AIs developed after an initial phase of suppression of T [T is converted to 17β-estradiol (E2) by aromatase]-dependent cell proliferation and survival.…”

“…So far, most studies focus on LTED-I, not LTED-H. Our AI-resistant model provides an opportunity to study ERα hypersensitivity, because these acquired AI-resistant cells grow in the presence of T and AI (a small amount of E2 can be converted in this situation) and are sensitive to fulvestrant (16). LTED cells do not respond to any AI, whereas our AI-resistant cells acquire resistance to one AI and will respond to another AI (16,32), which resembles a true clinical situation (33). Moreover, LTED cells are less responsive to fulvestrant compared with our AI-resistant cells (16), suggesting LTED cells are less dependent on ERα signaling than our resistant cell lines.…”