To define the function of SHBG and CBG, we've devised ways of modifying their expression in transgenic mice. We've developed a series of transgenic mice that express the human SHBG gene under the control of promoter sequences that are utilized in specific tissues, thereby presenting a unique opportunity to study the regulation of human SHBG in vivo and determine the functional significance of its transcripts in different tissues throughout normal development. We expect these studies should improve our understanding of steroid hormone bioavailability during development, and reveal new ways of manipulating steroid-dependent effects on cell differentiation and growth. These transgenic mouse models have also revealed that the production of human SHBG is regulated by nutritional factors, which we believe will allow us to demonstrate at the molecular level why plasma levels of SHBG are such a good of marker of metabolic status, and a prognostic indicator of risk for metabolic diseases, including type II diabetes and cardiovascular disease.

The expression of SHBG genes in other model organisms, including various fish species, is also a major focus of our research. Studies involving zebrafish have already revealed that the gut is an important site at which SHBG acts, and we believe that this is relevant to the functional role of SHBG in the developing gut of mammals as well. The central role of SHBG in regulating sex steroid action during development in fish is also a primary focus, especially in light of observations that small molecules in the environment are capable of interacting with the fish SHBG steroid-binding site, and may disrupt the binding of natural sex steroids. We believe that these types of compounds could have a major impact on reproductive development and reproductive performance in fish. A greater understanding of these effects could be relevant to the role(s) of SHBG in sexual development in mammals exposed to environmental contaminants, especially those that disturb the normal endocrine balance.

One of our major contributions over the past few years has been to generate a detailed picture of the structure of human SHBG, revealing for the first time exactly how various steroid ligands are accommodated within the SHBG steroid-binding site. We have also discovered that occupancy of a specific zinc-binding site within the human SHBG steroid-binding site can influence its affinity for estrogens. Currently, this structural information is being used to help us understand how SHBG interacts with other proteins, and how this is influenced by occupancy of the SHBG steroid-binding site. It has also allowed us to devise, in collaboration with Dr. Artem Chersakov, an in silico screening strategy for the identification of novel non-steroidal SHBG ligands. Our transgenic mouse models mimic the tissue and cell-specific expression profile of human SHBG, and provide the unique opportunity to evaluate the physiological effects of novel SHBG ligands identified through this powerful in silico screening strategy.

Through collaborations with clinical colleagues around the world, our work has led to the identification of several single nucleotide polymorphisms within the human CBG gene that result in the production of CBG variants with reduced affinities for cortisol. Individuals with these CBG gene variants have hypotension and fatigue, and their clinical profiles are similar to patients who have recently been shown by others to lack CBG in the blood due to a null mutation. Since the CBG gene is expressed in several tissues, including the kidney during late fetal and neonatal development, deficiencies in CBG gene expression during critical stages of development result in an imbalance of glucocorticoid levels that impact on the normal development of these tissues, and the appearance of disease in later life. Our efforts are now directed towards addressing these and other questions related to the physiological importance of CBG as a modulator of glucocorticoid action.

We've also identified several human SHBG gene variants with either a polymorphism in a regulatory element of the promoter or in the coding sequence, and at least one of these genetic variants is associated with hyper-androgenism and/or ovarian dysfunction in women. Further studies are in progress to assess whether these or other genetic variations in the human SHBG gene are associated with other disease processes linked with an imbalance in androgen or estrogen action.

Selva DM, Hammond GL (2009) Peroxisome-proliferator receptor represses hepatic sex hormone-binding globulin expression. Endocrinology Epub 2009 Jan 29.

Lin HY, Underhill C, Gardill BR, Muller YA, Hammond GL.: Residues in the human corticosteroid-binding globulin reactive center loop that influence steroid binding before and after elastase cleavage. J Biol Chem. 2009 Jan 9;284(2):884-96.

Pawluski JL, Charlier TD, Lieblich SE, Hammond GL, Galea LA.: Reproductive experience alters corticosterone and CBG levels in the rat dam. Physiol Behav. 2009 Jan 8;96(1):108-14.

Thorsteinson N, Ban F, Santos-Filho O, Tabaei SM, Miguel-Queralt S, Underhill C, Cherkasov A, Hammond GL.: In silico identification of anthropogenic chemicals as ligands of zebrafish sex hormone binding globulin. Toxicol Appl Pharmacol. 2009 Jan 1;234(1):47-57.

Miguel-Queralt S, Underhill C, Devlin RH, Hammond GL.: Characterization and measurement of the plasma alpha- and beta-sex hormone-binding globulin paralogs in salmon. Endocrinology. 2009 Jan;150(1):366-75.

Hammond GL, Hogeveen KN, Visser M, Coelingh Bennick HJ.: Estetrol does not bind sex hormone binding globulin or increase its production by human HepG2 cells. Climacteric. 2008;11(suppl 1):41-6.

Poon SL, An BS, So WK, Hammond GL, Leung PC.: Temporal recruitment of transcription factors at the 3',5'-cyclic adenosine 5'-monophosphate-response element of the human GnRH-II promoter. Endocrinology. 2008 Oct;149(10):5162-71.

Miguel-Queralt S, Hammond GL.: Sex hormone-binding globulin in fish gills is a portal for sex steroids breached by xenobiotics. Endocrinology. 2008 Sep;149(9):4269-75.

Cherkasov A, Ban F, Santos-Filho O, Thorsteinson N, Fallahi M, Hammond GL.: An updated steroid benchmark set and its application in the discovery of novel nanomolar ligands of sex hormone-binding globulin. J Med Chem. 2008 Apr 10;51(7):2047-56.

Selva DM, Hogeveen KN, Innis SM, Hammond GL (2007) Monosaccharide-induced lipogenesis regulates the human hepatic sex hormone-binding globulin gene. J Clin Invest 117:3979-87

Klieber MA, Underhill C, Hammond GL, Muller YA (2007) Corticosteroid-binding globulin, a structural basis for steroid transport and proteinase-triggered release. J Biol Chem 282:29594-603. Epub 2007 Jul 19.

Ng KM, Catalano MG, Pinos T, Selva DM, Avvakumov GV, Munell F, Hammond GL (2006) Evidence that fibulin family members contribute to the steroid-dependent extravascular sequestration of sex hormone-binding globulin. J Biol Chem 281:15853-61. Epub 2006 Apr 6.

Selva DM, Bassas L, Munell F, Mata A, Tekpetey F, Lewis JG, Hammond GL (2005) Human sperm sex hormone-binding globulin isoform: characterization and measurement by time-resolved fluorescence immunoassay. J Clin Endocrinol Metab 90:6275-82. Epub 2005 Aug 30.

Selva DM, Hogeveen KN, Hammond GL (2005) Repression of the human sex hormone-binding globulin gene in Sertoli cells by USF transcription factors. J Biol Chem 280:4462-4468

Miguel-Queralt S, Knowlton M, Avvakumov GV, Al-Nouno R, Kelly GM, Hammond GL (2004) Molecular and functional characterization of sex hormone binding globulin in zebrafish. Endocrinology 145:5221-52230

Selva DM, Hogeveen KN, Seguchi K, Tekpetey F, Hammond GL (2002) A human sex hormone-binding globulin isoform accumulate in the acrosome during spermatogenesis. J Biol Chem 277:45219-45225

Avvakumov GV, Grishkovskaya I, Muller YA, Hammond GL (2002) Crystal structure of human sex hormone-bindng globulin in complex with 2-methoxyestradiol reveals the molecular basis for high-affinity interactions with C2 derivatives of estradiol. J Biol Chem 277:45291-45298

Grishkovskaya I, Avvakumov GV, Hammond GL, Catalano MG, Muller YA (2002) Steroid ligands bind human sex hormone-binding globulin in specific orientations and produce distinct changes in protein conformation. J Biol Chem 277:32086-32093

Visiting Scholar Award, Sigrid Juselius Foundation, 2009
Tier I Canada Research Chair in Reproductive Health

Eui-Ju Hong, DVM - Graduate student
Hwa-Yong Lee, Postdoctoral fellow
Jason Popesku, Postdoctoral fellow
Gina Teodosio, Research technician
Caroline Underhill - Lab manager / research assistant / technician
Tsung-Sheng (John) Wu, Graduate student