Abstract 1602: Snail transcription factor regulates nuclear cathepsin L activity

Abstract

Abstract Triple negative breast cancer (TNBC) are defined as tumors lacking the expression of estrogen receptor-alpha, progesterone receptor and human epidermal growth factor receptor-2, which accounts for approximately 15% of total breast cancer patients, and is more prevalent and lethal among young African, African-American and Latino women patients. The primary cause of breast cancer death is metastasis, which is regulated by factors such as epithelial mesenchymal transition (EMT), a dynamic process that promotes cell motility and decreases cell-cell adhesion. Snail transcription factor promotes EMT by repressing epithelial genes such as E-cadherin, while increasing mesenchymal genes such as vimentin. Cathepsin L (Cat L) cysteine protease promotes cell invasion and nuclear Cat L has recently been associated with poor prognosis in colon and breast cancer. However, the mechanism by which Cat L localizes to the nucleus is unknown. CDP/CUX transcription factor is a substrate for Cat L which when proteolytically cleaved by Cat L from the p200 to the p110 isoform can transcriptionally activate Snail and repress E-cadherin. We have recently published that Snail overexpression increases Cat L activity in prostate cancer cells which can be antagonized by muscadine grape skin extract (MSKE), a natural product rich in anthocyanin. We hypothesize that a feedback loop exists whereby Snail Transcription factor may promote nuclear Cat L expression and activity in TNBC which subsequently cleaves CDP/CUX and further increases Snail transcription, leading to increased EMT; this can be abrogated by Cat L inhibitor or MSKE. We utilized various breast cancer cell lines as well as MCF-7 cells stably overexpressing Snail (MCF-7 Snail) or control (MCF-7 Neo) to evaluate the expression of Snail, Cat L, CDP/CUX and EMT marker expression (E-cadherin, vimentin) by western blot and immunofluorescence. Migration and invasion assays were performed including treatments with Cat L specific inhibitor (Z-FY-CHO) or MSKE. We observed more Snail, Cat L and CDP/CUX cleavage products (p110 and p90 isoforms) expression in MCF-7 Snail cells and TNBC cells (MDA-MB-231, MDA-MB-468) as compared to MCF-7 control cells. Interestingly, immunofluorescent and cell fractionation analyses revealed that Snail overexpression promoted nuclear Cat L and CDP/CUX p110 isoform expression, and EMT (low E-cadherin, high vimentin, migration and invasion) which could be abrogated by Z-FY-CHO or MSKE. We are currently staining for Snail and Cat L in breast cancer tissue of varying races/grades including TNBC, as well as confirming by chromatin immunoprecipitation that the CDP/CUX cleavage products seen in TNBC can bind Snail and/or E-cadherin promoters. In conclusion, our study shows that Snail promotes nuclear localization of Cat L which may promote EMT via CDP/CUX, and that inhibition with Cat L inhibitors or natural products such as MSKE may be a good therapeutic target for TNBC. Citation Format: Liza J. Burton, Valerie Odero-Marah. Snail transcription factor regulates nuclear cathepsin L activity. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1602.