A Wind of Change with Wnt

The diabetic population is constantly increasing. In 2010, an estimated 285 million people worldwide were affected by diabetes mellitus, 90% of whom had the type 2 form. Type 2 diabetes mellitus (or adult-onset diabetes) is characterized by hyperglycemia due to insulin resistance, inadequate insulin secretion and excessive or inappropriate glucagon production. It can become life-threatening when left untreated and is associated with environmental risk factors such as age, sex, obesity, low physical exercise, smoking, diet and stress. The latter are thought to reduce insulin release and activity in individuals with a genetic susceptibility to the disease.

The maintenance of blood glucose homeostasis relies on the coordination of several physiological systems including the sympathetic nervous system and the endocrine system. More recently, Wnt signaling has been tightly linked with diabetes following the identification of mutations in one of its major effector, the transcription factor TCF7L2, and the increase of metabolic regulators like incretin, insulin, pro-glucagon and GLP-1. In the canonical Wnt signaling pathway, Wnt binds to Frizzled and LRP5/6 co-receptors and triggers a signaling cascade that stabilizes β-catenin. The latter translocates from the cytoplasm into the nucleus and induces transcriptional activation of target genes. Several naturally occurring antagonists are capable of regulating Wnt signaling in vivo. For instance, Dickkopf (DKK) proteins can bind to the LRP5/6 co-receptors and prevent them from interacting with Wnt-Frizzled complexes. Professor Wu and colleagues from Yale School of Medicine collaborated with Enzo Therapeutics and investigated the involvement of the antagonist DKK2 in the regulation of glucose metabolism by means of computational and biological screening of small molecule inhibitors as well as murine genetic approach. Using Enzo’s LeadingLight® Wnt reporter assay kit, fifty-four compounds were screened for their ability to reverse DKK-mediated inhibition of Wnt signaling. This assay is a cell-based luciferase activity test, which uses an engineered cell line expressing the firefly luciferase reporter gene under the control of Wnt-responsive promoters (TCF/LEF). The luciferase activity from the reporter gene in this cell line is regulated in a dose-dependent manner upon the addition of exogenous Wnt or related ligands, agonists or antagonists to the cell culture medium.

The DKK inhibitor IIIC3 (gallocyanine) gave the most significant and reproducible results with efficient and potent reversal of DKK inhibition. IIIC3 and several custom-made analogs were then further characterized. Both IIIC3 and the analog IIIC3a were found to directly bind to LRP5/6 and compete with DKK-LRP5/6 interaction. Mice treated with either IIIC3 or IIIC3a showed significantly lower blood glucose concentration and improved glucose tolerance without altering insulin production or sensitivity. The same phenotype was observed in Dkk2-knockout mice suggesting that the positive effects on glucose metabolism of IIIC3 and IIIC3a were obtained by antagonizing DKK2. Finally, intra-peritoneal administration of IIIC3a to a murine model of type 2 diabetes led to a significant decrease in blood glucose concentration and a better tolerance to glucose. Altogether, these results suggest that Wnt signaling and more specifically DKK2 represents an interesting therapeutic target for treating type 2 diabetes mellitus. It remains to be seen, however, if an increase in Wnt activity, even moderate, does not pose a threat by increasing the risk of developing cancer.

Enzo Life Sciences offers a comprehensive portfolio for studying glucose metabolism and Wnt signaling including cell-based reporter assay, ELISA kits, antibodies, proteins and biochemicals; some of which are listed below:

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