Abstract 4942: Activation of Phosphatase toward the Retinoblastoma protein in breast and colorectal cancer cell spheroids

Abstract

Abstract Excessive phosphorylation of the Retinoblastoma protein is found in most cancer tumor types. Several cyclin dependent kinase (cdk) inhibitors are in development or in clinical trials at the present time. Our previous studies have focused on the regulation of Rb phosphorylation by the phosphatase, PP1. Specificity toward substrates is imparted onto PP1 by many different interacting proteins. In proliferating cells, PP1 is associated with a regulatory protein called PNUTS (Phosphatase Nuclear Targeting Subunit). Our previous experiments have shown that PNUTS inhibits PP1 activity toward specific Rb phosphorylation sites. Further we have demonstrated that siRNA mediated knockdown of PNUTS in several cancer cell types (colon, breast, ovarian) leads to an increase in Rb specific PP1 activity, Rb dephosphorylation on several sites, and a dramatic induction of apoptosis. Furthermore, the ability of PNUTS siRNA to induce apoptosis was shown to be dependent on Rb expression. Interestingly, further studies showed that PNUTS knockdown had no effect on non-transformed breast and colorectal cells, in which Rb is not hyperphosphorylated. Thus we have developed a method to activate the Rb phosphatase in cells by knockdown of the PP1 binding protein, PNUTS. The current study was undertaken to analyze the effect of PNUTS knockdown in cancer cells grown in three-dimensional spheroids grown by incubation of cells in hanging drop plates. The effect of PNUTS knockdown on Rb phosphorylation, cell proliferation and cell viability using this model system will be presented. Citation Format: Jacklynn Egger, Lisa Antonucci, Maria Lane, Nancy A. Krucher. Activation of Phosphatase toward the Retinoblastoma protein in breast and colorectal cancer cell spheroids. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4942. doi:10.1158/1538-7445.AM2015-4942