In breast cancer, low retinoic acid-2 (RAI2) gene expression is correlated with presence of disseminated tumor cells in the bone marrow of early stage patients and with poor outcome. Furthermore, the RAI2 protein acts as a transcriptional co-regulator involved in hormonal response and differentiation of hormone dependent breast cancer cells. Here, we evaluate the role of RAI2 in prostate and breast cancer, both steroid hormone driven cancers, focusing on its impact on progression to a hormone independent disease.
First protein expression of RAI2 and hormone receptors (HR) were analyzed in hormone dependent breast and prostate cancer cell lines after treatment with RAI2- or HRspecific shRNAs. Quantitative PCR was applied to examine expression of HR regulated genes in RAI2-depleted VCaP prostate cancer cells and KPL-1 breast cancer cells after hormone stimulation. The effect of a somatic RAI2 knockout in LNCaP prostate cancer cells and KPL-1 breast cancer cells on hormone independent growth was analyzed under hormone deprivation or HR inhibition. Anchorage-independent growth as well as cellular motility of RAI2 knockout LNCaP cells were also tested. Protein expression of the MAPK/ERK and PI3K/AKT signaling axis was analyzed in RAI2 knockout KPL-1 cells, cultured under hormone deprivation or fulvestrant treatment.
Silencing of RAI2 decreased the expression of AR in prostate cancer and ER in breast cancer cell lines, respectively. Moreover, silencing of hormone receptors increased RAI2 expression in cell lines of both tumor entities, revealing an interdependent regulation of RAI2 and tested hormone receptors. Furthermore, after hormone stimulation RAI2 depleted VCaP cells showed increased induction of androgen target genes like STEAP4, MME and SGK1. Similarly, loss of RAI2 expression in KPL-1 cells increased the induction of several estrogen target genes like EGR3, PGR and SNAI1 after hormone stimulation. In LNCaP prostate cancer cells complete loss of RAI2 significantly increased cell viability, colony formation, anchorage independent growth and cell migration. Additionally, those cells were more resistant to chemical castration. Similarly, KPL-1 cells were less sensitive to anti-estrogens and showed an additional growth advantage in hormone free media after RAI2 loss. Analysis of differentially expressed proteins revealed an upregulation of HER2 in RAI2 knockout KPL-1 cells under hormone deprivation.
Our results show that RAI2 acts as a corepressor of steroid regulated genes in prostate and breast cancer cells. Moreover, we revealed a functional association between RAI2 and hormone dependent tumor growth. RAI2 loss could thus contribute to resistance to endocrine therapy in both cancer entities by the induction of receptor tyrosine kinases, like HER2.
Citation Format: Katharina Besler, Aleksandra Weglarz, Christina Zill, Jana Jensen, Harriet Wikman, Klaus Pantel, Stefan Werner. Role of RAI2 protein in progression of breast and prostate cancer to hormone independent disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3736.