Abstract 2644: Inhibition of choline kinase increases endoplasmic reticulum stress proteins
Authors: Sanchez-Lopez Elsa , Menchén Laura , Seco Esther , del Pulgar Teresa Gómez , Lacal Juan Carlos , Cebrián Arancha 
Published: June 2012

Abstract
Choline kinase (ChoK), the first enzyme in the Kennedy pathway, is responsible for the generation of phosphorylcholine (PCho) that renders phosphatidylcholine, the most abundant phospholipid of plasma membrane. ChoK overexpression has been detected in several cancer types and may act as a prognostic factor in lung, bladder and breast tumors, suggesting its important role in proliferation and cell growth. Specific pharmacologic or siRNA ChoKα inhibition have been proposed as a novel broad-spectrum antitumor strategy. Indeed, ChoKα inhibition induces cell death in several tumor cell lines but is a cytostatic drug in primary cells. In order to exploit their full potential as novel anticancer drugs, it is essential to understand the mechanism implicated in the differential cytotoxic effects induced by ChoK inhibitors on cancer cells. Here we investigate this mechanism using as a model cells from a ductal mammary carcinoma, T47D, and the non-tumorigenic epithelial gland mammary cells, MCF10A.
Cells were treated with two chemical ChoK inhibitors, MN58b and TCD717, the latter recently approved by the FDA to begin a Phase I clinical study for the treatment of solid tumors (http://clinicaltrials.gov/ct2/show/NCT01215864). First, cell viability and proliferation was evaluated to confirm the previous differential effect described in other cell types. By flow cytometry and MTT assay both inhibitors induced cell death in the T47D cells but caused G0/G1 cell cycle arrest in the non-tumorigenic MCF10A cells. Protein levels of several cell cycle (cyclins, phosphor-Rb, E2F) and cell death related proteins (caspase3, PARP) were studied, confirming that ChoK inhibitors caused cell cycle arrest in normal cells but cell death in breast cancer cells. To further investigate the mechanism implicated in this specific cytotoxic effect, the expression levels of some of the most relevant ER stress related genes such as C/EBPβ, ATF4 and CHOP were investigated by QPCR. These genes have been recently involved in apoptosis induced by cytotoxic agents such as tocotrienol and delta(9)-tetrahydrocannabinol. C/EBPβ gene expression levels were significantly increased in tumor cells treated with either inhibitor compared to the non-tumourigenic cells, and this increase was maintained from at least 6 to 48 hours. CHOP overexpression was also significantly increased at earlier times after treatment in T47D cells than in MCF10A cells. Moreover, treatment with ChoK inhibitors significantly increased the production of CHOP and the major ER chaperone BIP in T47D cells compared with non-tumourigenic cells. These data suggest a differential effect of ChoK inhibitors on the induction of ER stress factors in normal vs. tumor cells.
In summary, ChoK inhibition has a profound differential effect in cell cycle regulation and in ER stress response in tumor vs. normal cells, suggesting that ER stress could mediate the specific cytotoxic effects of ChoK inhibitors.
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 2644. doi:10.1158/1538-7445.AM2011-2644