Abstract 2348: Cyr61 and FOXOA1 are inversely regulated in IGF-1 mediated breast cancer cells
Authors: Sarkissyan Suren , Sarkissyan Marianna , Wu Yanyuan , Koeffler H. Phillip , Vadgama Jaydutt V. 
Published: June 2012

Background: Studies from our laboratory and others have shown that cysteine-rich 61 (Cyr61), a protein involved in wound healing and regulation of extracellular matrix may be involved in tumor proliferation and invasion. The mechanisms that regulate Cyr61 in aggressive tumors are not well understood. We have shown that IGF-I mediated PI3K/Akt activation results in increased Cyr61. Recently, we showed that FOXO1A, a member of the FKHR transcription family plays an important role in drug resistance. FOXO1A is a downstream target of the PI3K/Akt pathway. In this study, we will investigate mechanisms that regulate IGF-1 induced Cyr61 activation with transcriptional downregulation of FOXO1A.

Methods: Several breast cancer cell lines were used in this study: MDA-MB231, BT474, MCF-7, SKBR3, and MCF-7 CYA. Also tested were cells stably transfected with active Akt1 (SKBR3/AA and MCF-7/AA) and dominant negative Akt1 (SKBR3/DN and MCF-7/DN). The MCF-7 CYA cell line is stably transfected with a vector over-expressing Cyr61. Monolayer cultures were treated with IGF-I and Cyr61 expression was analyzed by Real-time PCR. Migration assays were used to determine IGF-1/Cyr61 activated cell invasion. Sphere assays were used to assess adherence-independent growth of tumor cells. Cyr61 and FOXO1A levels in wild type and CYA cells were assessed using immunofluorescent staining. The PI3K/Akt inhibitor, LY294002 was used to determine the PI3K/Akt dependent interactions between Cyr61 and FOXO1A.

Results: Our result show that cells with overactive pAkt, as in SKBR3/AA and MCF-7/AA, have increased levels of Cyr61. Conversely, cells with inactive Akt, such as in SKBR3/DN and MCF-7/DN have decreased levels of Cyr61. IGF-I treatment increased Cyr61 expression significantly and cells with high level of Cyr61 demonstrate increased invasiveness. Cyr61 over-expressing cells are more likely to form spheres and in greater numbers than cells with low Cyr61. Furthermore, Cyr61 over-expressing cells (MCF-7 CYA) have decreased FOXO1A. IGF-1 caused a greater downregulation of FOXO1A in MCF-7 CYA cells compared to wtMCF-7 cells. This down regulation of FOXO1A was restored to normal levels upon treatment with PI3K/Akt inhibitor LY294004 in wtMCF-7 but not in MCF-7 CYA cells. These data suggest that FOXO1A is constitutively downregulated in Cyr61 overexpressing cells and these cells when exposed to growth factors such as IGF-1, show further loss of FOXO1A

Conclusion: Our study demonstrates an inverse relationship between Cyr61 and FOXO1A expression in breast cancer cells. Cells with high Cyr61 and low FOXO1A are more invasive. IGF-I activates pAkt, increases Cyr61 and decreases FOXO1A expression. The PI3K inhibitor LY is able to restore FOXO1A levels in the presence of IGF-1 in MCF-7 cells, but not in MCF-7 cells with high Cyr61. Hence, targeting FOXOA1 in breast tumors which have high Cyr61 and are exposed to growth factors may offer a therapeutic potential.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2348. doi:10.1158/1538-7445.AM2011-2348