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Effective Screening Systems for the Discovery of Hormone-Like Therapeutic Compounds
David W. Wood
([email protected])
Department of Chemical Engineering
Background - Approximately 4% of the drug targets of current therapies belong to the nuclear hormone
receptor superfamily. These transcription factors control the expression of several genes in response to
the presence of small-molecule hormones or hormone-like compounds, and include the estrogen,
androgen, thyroid hormone, progesterone and vitamin D receptors among others. Their function has been
linked to a broad spectrum of diseases, including breast, endometrial and prostate cancer, leukemia,
cardiovascular diseases, osteoporosis and inflammations. Therefore the discovery of novel compounds
with the ability to modulate these targets could lead to the development of valuable therapeutics against
serious pathological conditions. Typical methods for identifying these compounds have included
engineered in vivo reporter systems, as well as in vitro receptor binding assays. More recently, small
peptides which bind and modulate receptor activity have been identified using combinatorial methods.
Description - In our previous work work, we developed a novel sensor of nuclear hormone binding in
Escherichia coli by constructing a gene fusion that combined the ligand binding domain of the α-subtype
of the human estrogen receptor with a thymidylate synthase enzyme (TS). Expression of this fusion in
TS-deficient bacterial cells resulted in estrogen-dependent cell growth. Subsequent replacement of the
estrogen receptor with the ligand-binding domain of the human thyroid hormone receptor led to thyroid
hormone-dependent growth that was insensitive to estrogen. Remarkably, this simple biosensor is able to
discriminate between agonistic and antagonistic activities, as combinations of estrogen agonists had an
additive impact on cell growth, whereas known estrogen antagonists were found to neutralize agonist
effects. More recently we have inserted the β-subtype of the human estrogen receptor, and have used this
construct to detect estrogenic compounds with high reliability and sensitivity. We have further been able
to detect compounds with subtype-selective activity, as well as a wide range of environmental and
pollutant estrogens1. Most significantly, our system was the first to identify three new estrogenic
compounds from a recently synthesize small-molecule library, and estrogenicity of all three of the
compounds have new been confirmed in human breast cancer and endometrial cells. Estrogen
antagonists are particularly important in the treatment of breast cancer, and include such compounds as
Tamoxifen and Raloxifene.
Commercial Potential - This is a practical technology for screening large libraries of compounds for the
ability to modulate the activity of specific human nuclear hormone receptors. These libraries might
include exogenous small-molecule compounds, as well as evolved peptides. The system is also designed
to work flexibly with a large number of hormone receptors, including underdeveloped targets and poorly
characterized orphan receptors. This work has recently led to ongoing discussions on optimization and
adaptation of the system for use with very large compound libraries and proprietary high-throughput
screening technologies. It is anticipated that these collaborations will yield significant improvements,
allowing it to be used effectively for drug discovery and identification.
Status and Future Plans - Thus this system has proven utility in the screening of a small library of novel
compounds for potential therapeutics, and has been disclosed and a patent applied for by Princeton
University. The appeal of this system will increase with the development of accessory technologies that
apply our sensor to the rapid identification or evolution of potentially therapeutic compounds.