Research in the Guy laboratory focuses on two main areas: assembly of the transcriptional activation complex by nuclear hormone receptors, especially the thyroid, androgen, and glucocorticoid receptors; and, the development of novel therapeutic agents for pediatric orphan diseases, especially malaria, retinoblastoma, ependymoma, and sleeping sickness. Our major technique focuses are parallel synthetic and medicinal chemistry, high content screening, and high throughput screening.
Intracellular signaling pathways depend upon careful modulation of protein interactions for faithful transmission of signal from origin to effector. One major research focus of the laboratory is the dissection of these interactions, which often involve the potential interaction of one family of proteins with another but paradoxically provide for precise selectivity in vivo. Our major focus has been understanding the regulation of transcription by the nuclear hormone receptors (NR's). The NR's are a superfamily of DNA binding proteins that regulate target gene transcription in response to circulating non-peptide hormones. NR's exert their effects by nucleating the formation of a multiprotein regulatory complex with many accessory proteins including coactivators, corepressors, and the basal transcription machinery.
Quantitative Proteomics of the NR
Often critical protein interactions in signaling involve the interaction of one family of factors with another. In this context, simply identifying the relevant players can be a significant challenge. The NR's represent such a case. They are structurally dynamic proteins that adopt discrete conformations in the unliganded state or when bound to an agonist or antagonist ligand. These changes in conformation alter the interactions with other proteins in the complex, particularly coactivators and corepressors and thereby induce the different signaling effects of agonist and antagonist. Unfortunately, a precise understanding of the mechanisms by which the receptors exert their effects is still lacking. We have developed a set of high throughput assays that allow determination of which co-regulatory protein is recruited to a given receptor in a particular state. As these systems have matured, we have begun to develop microarray-based methods for determining which response elements are activated in situ in order to allow full study of the systems involved. The goal is to eventually be able to map the circuitry underlying hormonal signaling using inferential modeling methods.
Proteomimetic inhibitors of interactions
The highly dynamic protein interactions in signaling pathways are often mediated by small contact areas of highly defined structure - usually common secondary structures such as turns, helicies, or strands. We have focused upon the design and synthesis of generally useful mimetics of such protein secondary structures and the evolution of these general scaffolds towards high affinity and selectivity for particular protein interactions. We use a two-phase strategy, beginning with a constrained peptide and following up in successful systems with a designed small molecule peptidomimetic. Our first approach to alpha helical proteomimetics has delivered useful and highly selective inhibitors of the interaction of the estrogen and thyroid receptors with their co-regulatory proteins. We have also developed the first small molecule inhibitors of the interaction of the thyroid receptor and androgen receptor with their obligate co-factors. An important underlying theme in this program is experimental validation of particular computational design methods and intensive use of parallel chemical synthetic techniques that inherently drives advances in library synthesis methodologies.
In the last five years, a new research direction has developed within the Guy group: the discovery of new therapeutic leads for orphan diseases. A number of diseases that cause a significant burden upon worldwide human health do not enjoy the benefits of industrially targeted drug development - predominantly due to unfavorable market analysis on the part of pharmaceutical companies. For the most part, these diseases are characterized as either being an orphan disease, one in which there are fewer than 200,000 patients worldwide, or as a disease of the developing world. Both of these conditions lead to unprofitable market: the former through small numbers of patients, the latter through their inability to pay for treatment.
Our program is aimed at producing candidate compounds for several of these diseases: malaria, retinoblastomas, ependymomas, and sleeping sickness. These efforts are being carried out in the context of a consortium of investigators who are deliberately pooling our efforts to bridge the "translation gap" between target identification and completion of pre-clinical studies of potential new therapies. Our group has focused on applying high throughput chemistry and high throughput screening methods to target discovery, target validation, lead discovery, and lead optimization. Our general strategy is to use a scaffold based chemical approach to produce highly diverse screening libraries based upon "privileged scaffolds," to screen these libraries for lead compounds, and to follow up the leads through focused library production and screening. Important current projects are the optimization of novel quinolines that are active against drug resistant strains of Plasmodium falciparum, the optimization of a protease inhibitor to provide compounds active against Trypanasoma brucei, and the discovery and optimization of novel inhibitors of the interaction of p53 with MDM2 and MDM4.
The majority of our projects depend upon the production and use of chemical libraries. Our work centers on the synthesis, purification, and analysis of high purity parallel libraries of hundreds to thousands of compounds. We also have interests in the storage and use of larger screening libraries including defining high quality libraries and the automated analysis of HTS data and generation of predictive models of structure activity relationships from that data. Finally, we work on the development of new methods for high information content screening (HCS).