The Ras oncogene has been known as a key player in development of the majority of human tumors for over 25 years.  Its function has turned out to be deceptively complex and many aspects of its biology remain poorly understood.  In part, this is because Ras binds a multitude of heterologous effectors to control a complex, interconnected signaling network.  Although much attention has been focused on the ability of Ras to activate the MAP kinase pathway via Raf proteins, the ability of Ras to activate non-Raf effectors may be more important to its role in human cancer.  In particular, we are interested in two of the relatively obscure Ras signaling pathways mediated by RASSF family proteins and by the RalGEF family.

RASSF proteins appear to serve as pro-apoptotic effectors of Ras that are down-regulated in human tumors to permit dangerously excessive Ras activity to drive transformation without inducing cell suicide by apoptosis.  RalGEF proteins are a group of Ras effectors that can connect Ras to the activation of Ral proteins.  The role of RalGEF in transformation remains somewhat obscure and activated forms of RalGEF are poorly transforming in murine systems.  However, more recent work has identified RalGEFs as key components of Ras transformation in human cells.

RASSF proteins are usually inactivated in tumors by an epigenetic process involving aberrant promoter methylation leading to transcriptional silencing.  Thus, the gene remains intact in the tumor cell, just dormant.  Treatment of cells with inhibitors of the DNA Methyl Transferase (DNMT) enzyme system can restore expression of RASSF proteins.  However, most of these drugs are very toxic, which may limit their utility.  A new DNMT inhibitor has recently been described (RG108) with a different mechanism of action and a much lower toxicity.  We have now found that it can reactivate the expression of the RASSF1A protein in tumor cells.  As RG108 has low toxicity, we may be able to use it in combination with other inhibitors of Ras action to synergistically inhibit Ras-dependent transformation..

The long-term goals of the project are to:

1. Understand the mechanism of action of RASSF proteins to enhance our ability to use their inactivation as a prognostic and diagnostic tool as well as a novel therapeutic target
2. Develop novel inhibitors of a key Ras signaling pathway, RalGEF; and
3. Determine if using small molecules to reactivate RASSF inhibitory pathways and at the same time using novel small molecules to inactivate the transforming RalGEF pathway we can generate proof of principal of a novel synergistic inhibition of Ras-mediated transformation.