Abstract B36: RENCA macrobead-induced AKT hyperphosphorylation leads to MEF2 activation and inhibition of the proliferation of human DU145 prostate carcinoma cells
Authors: Martis Prithy C. , Laramore Melissa A. , Dudley Atira T. , Smith Barry H. , Gazda Lawrence S. 
Published: May 2016

We have previously reported that suppression of the isoforms of myocyte enhancer factor (MEF2) in target tumor cells in vitro reduces the demonstrated proliferation-inhibitory effect of agarose-agarose encapsulated mouse renal adenocarcinoma cells (RENCA macrobeads), suggesting that it is a critical component of a pathway by which the inhibitory action, now being evaluated also in Phase IIb human clinical trials, is mediated. Importantly, MEF2 originally described as active in myogenesis, has been found to be a key regulator of many developmental pathways and to have both pro- and anti-growth regulatory functions in murine and human tumor cells.

Recent studies have supported a molecular intersection between components of the PI3K/Akt and the canonical Wnt signaling (carcinogenic) pathways, converging at the glycogen synthase kinase 3β (GSK3β) node. In skeletal and cardiac muscle cells, GSK3β regulates MEF2 activity indirectly through reciprocal regulation of the p38 mitogen-activated protein kinase (MAPK) pathway.

To determine, therefore, whether PI3K/Akt is also involved in the anti-proliferative effect of the RENCA macrobeads, we evaluated signaling through PI3K/Akt in response to macrobead-conditioned media and examined post-translational modification of MEF2D in two human cell lines DU145 (prostate cancer) and MCF7 (breast adenocarcinoma). Using In-Cell Western analysis, we determined the expression of phospho-Akt T308 and S473 in parallel with total Akt in cells exposed to naïve or conditioned media. Nuclear extracts for phospho-MEF2D and total MEF2D were assessed using western blotting. In the DU145 cells, exposure to RENCA macrobead-conditioned medium resulted in Akt hyperphosphorylation (≥ 48 hours) in parallel with the increasing age and inhibitory capacity of macrobeads. Further, the growth inhibition of DU145 cells exposed to RENCA conditioned medium (36-40% at 12-16 weeks of age; 45-48% at 20-24 weeks of age) was accompanied by de-phosphorylation of MEF2D at Serine 444, supporting a role for increased MEF2D transcriptional activity in the inhibitory effect. In the MCF7 cells, which are less sensitive to RENCA macrobead inhibition (8-12% at 12-16 weeks of age; 17-21% at 20-24 weeks of age), Akt phosphorylation diminished one hour following exposure to conditioned media, and MEF2D remained phosphorylated at Serine 444.

These findings support the hypothesis that RENCA macrobeads, as biological cell systems with the in vitro and in vivo (model and clinical) capability of inhibiting cancer cell proliferation and tumor growth, achieve this effect, at least in part, by signaling through Akt to differentially regulate MEF2D.

Citation Format: Prithy C. Martis, Melissa A. Laramore, Atira T. Dudley, Barry H. Smith, Lawrence S. Gazda. RENCA macrobead-induced AKT hyperphosphorylation leads to MEF2 activation and inhibition of the proliferation of human DU145 prostate carcinoma cells. [abstract]. In: Proceedings of the AACR Special Conference: Developmental Biology and Cancer; Nov 30-Dec 3, 2015; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(4_Suppl):Abstract nr B36.