SBIR-STTR Award

The production of adrenal gland hormones, including cortisol and androgens, is regulated by pituitary ACTH (adrenocorticotropin hormone), which acts via the melanocortin-2 receptor (MC2R) to stimulate steroid production. The ability to antagonize ACTH action would greatly facilitate the care of several clinical conditions, including congenital adrenal hyperplasia and Cushings Disease, which affect the adrenal gland. Yet, ACTH antagonists are not currently available. Thus the specific aim of this proposal is to discover new small molecule blockers of the MC2R with the long-term goal to develop them into drugs to treat the diseases mentioned above. For the purpose of this proposal, we will use Artificial Neural Networks and Support Vector Machines classification methods to design and synthesize targeted drug discovery library of potential ligands for peptide binding G-protein coupled receptors (GPCR), like MC2R. As a screening assay system, we have developed a dual expression sensor cell line, which simultaneously expresses MC2R and GFP-beta-Arrestin fusion protein. Upon the receptor stimulation with ACTH, the MC2R-GFP-beta-Arrestin complex migrates into endosomes. The process can be visualized and quantified with a fluorescent microscopy. In the presence of the receptor inhibition, we expect that the MC2R-GFP-beta-Arrestin complex will be prevented, which will allow for initial hit selection. The hits found will be entered into follow up investigation using both receptor binding techniques to characterize their affinities on membrane preparations containing the MC2R and functional cell responses, cAMP accumulation, to characterize their potencies and efficacies. These studies will involve the combined efforts of Chemical Diversity Labs, Inc., which has expertise in drug discovery chemistry, and a Yale University laboratory with expertise in GPCR action and the treatment of adrenal disorders.

Thesaurus Terms:
adrenocorticotropic hormone, drug discovery /isolation, hormone inhibitor, peptide library, small molecule adrenal disorder, arrestin, computer assisted sequence analysis, computer simulation, computer system design /evaluation, cyclic AMP, hormone receptor, molecular biology information system, receptor binding biotechnology, green fluorescent protein, microarray technology, molecular cloning, tissue /cell culture, transfection /expression vector

Development of drugs based on the MC2R antagonists will directly benefit children with CAH and Cushing's diseases, and will help us to understand MC2R's role in metabolic pathways. The long term goal of this project is to discover small molecule inhibitors of MC2 receptor and develop drug to treat several metabolic diseases including CAH and Cushing's diseases afflicting children. Our Phase I studies to identify novel ACTH small molecule antagonists were successful; we were able to identified three novel non-peptide small molecule ACTH antagonists that have high affinity to the MC2R. The inhibitors belong to the same chemotype with a similar chemical backbone. These compounds are full ACTH antagonists that potently inhibit ACTH-stimulated adrenal cell activity. We have achieved the Phase I goals and discovered a novel series of small molecule non-peptide ACTH antagonists. Phase II of this project will be dedicated to extending our studies of ACTH antagonists by examining structure-activity relationships using series analogs of identified inhibitors to assess their ACTH antagonist potency, specificity, and selectivity, and initiating animal testing. Specifically we will: 1) examine ACTH antagonist structure-activity relationships to increase our understanding of basic structural requirements for the compound to bind to MC2R; 2) assess selectivity and specificity of the ACTH antagonists to other melanocortin and unrelated receptors; (3) assess ADME- related characteristics such as human serum albumin binding, P450 inhibition, lipid membrane, CACO-2 permeability, and in vitro cytotoxicity; 4) perform in vivo animal toxicology/pharmacokinetic testing; and 5) assess ability to block ACTH action in an in vivo animal model. These studies combine ChemDiv's expertise in drug discovery with a Yale laboratory with expertise in GPCR action and treatment of adrenal disorders. The studies will lead to novel approaches for treating patients with ACTH-dependent adrenal disorders.

Project narrative:
The production of adrenal gland hormones, including cortisol and androgens, is regulated by pituitary ACTH (adrenocorticotropin hormone), which acts via the melanocortin-2 receptor (MC2R) to stimulate steroid production. Serious clinical disorders affecting the adrenal gland include congenital virilizing adrenal hyperplasia (CAH) and Cushing's disease. The ability to antagonize ACTH action will greatly facilitate the care of these and other conditions affecting the adrenal gland. Unfortunately, high-affinity small molecule ACTH antagonists are not identified yet and are not currently available for clinical use.

Thesaurus Terms:
adrenocorticotropic hormone, drug discovery /isolation, hormone inhibitor, peptide library, small molecule adrenal disorder, arrestin, computer assisted sequence analysis, computer simulation, computer system design /evaluation, cyclic AMP, hormone receptor, molecular biology information system, receptor binding biotechnology, green fluorescent protein, microarray technology, molecular cloning, tissue /cell culture, transfection /expression vector