Background and hypothesis: While grp94 expression in cancer is linked to aggressive disease and resistance to therapy, little is known on mechanisms underlying these effects. Our inability to study grp94 in genetically tractable organisms such as yeast and the unavailability of small molecule grp94 inhibitors are largely responsible for this state of affairs. Until now investigation of grp94 has mainly used mutant cell lines and gene deficient mouse studies. Although powerful, this approach is limited because it addresses phenotypic changes in the complete absence of a gene, and moreover, makes use of an engineered cellular environment. Alternative strategies that address the role and biology of grp94 in an endogenous cellular environment, where grp94 is limiting but not absent, are therefore needed. In addition to linking grp94-mediated mechanisms to the relevant disease state, i.e. tumor type, these reagents may provide evidence refractory to the currently available means. Approach: Discovery of grp94-selective ligands and chemical tools has been challenging thus far because of a high degree of analogy between the HSP90 paralogs. We here provide preliminary evidence showing that the discovery of grp94 inhibitors with log selectivity over the other paralogs is possible. Using a combination of screening an in-house generated small-molecule library and computational analyses backed by structural studies conducted with Project 3, we have identified important structural determinants that impart ligand selectivity and high affinity for grp94. We propose to use this information as a springboard for the development of grp94-directed chemical tools such as grp94 selective ligands and solid support immobilized, biotinylated and fluorescently labeled derivatives (Aim 1a) and as a starting point towards the development of drug-like grp94 inhibitors for translation to clinic as novel anti-cancer therapeutics (Aim 1b). We also propose here to use the grp94-directed toolset for the investigation of disease-specific roles of grp94 (Aim 2). Significance: In support of this investigative chemical biology approach, we provide preliminary data that confirm its power. We show that it identifies a novel role for grp94 in regulating HER2 tyrosine kinase at the plasma membrane, specifically in the case of tumors with overexpression of this protein. Also we implicate grp94 in regulating oncogenic signal transduction at the plasma membrane, indicating grp94 as a target in HER2-overexpressing breast cancer. These findings provide the blueprint for the development of grp94 inhibitors as a novel targeted therapy for the treatment of breast cancers dependent on increased signaling through plasma membrane receptors. Importantly, although HER2 was for the last two decades one of the most widely studied HSP90 onco-client protein, the mechanistic and the therapeutic understanding unveiled by our chemical biology approach was missed so far, proving further evidence to the importance of having a strong chemical toolset in addition to genetic and biochemical means for the understanding of biology.
This project will address an unmet research and medical need by developing novel chemical tools to enable a spatiotemporal investigation of grp94-regulated cancer mechanisms in endogenous cancer phenotypes. This chemical biology/pharmacology approach complements the genetic and structural approaches of other research components in this integrated program. Importantly, it will help address the biology of grp94 in the cellular environment where grp94 is limited but not absent, and this also aims to improve the drug-like characteristics of grp94 ligands towards the ultimate goal of identifying leads with therapeutic potential for clinical translation.
Showing the most recent 10 out of 55 publications
