The steroid hormone estrogen regulates many functionally unrelated processes in numerous tissues. Although it is traditionally thought to control transcriptional activation through the classical nuclear estrogen receptors, it also initiates many rapid nongenomic signaling events. We found that of all G protein-coupled receptors characterized to date, GPR30 is uniquely localized to the endoplasmic reticulum, where it specifically binds estrogen and fluorescent estrogen derivatives. Activating GPR30 by estrogen resulted in intracellular calcium mobilization and synthesis of phosphatidylinositol 3,4,5-trisphosphate in the nucleus. Thus, GPR30 represents an intracellular transmembrane estrogen receptor that may contribute to normal estrogen physiology as well as pathophysiology.

Estrogen is a hormone critical in the development, normal physiology and pathophysiology of numerous human tissues. The effects of estrogen have traditionally been solely ascribed to estrogen receptor alpha (ERalpha) and more recently ERbeta, members of the soluble, nuclear ligand-activated family of transcription factors. We have recently shown that the seven-transmembrane G protein-coupled receptor GPR30 binds estrogen with high affinity and resides in the endoplasmic reticulum, where it activates multiple intracellular signaling pathways. To differentiate between the functions of ERalpha or ERbeta and GPR30, we used a combination of virtual and biomolecular screening to isolate compounds that selectively bind to GPR30. Here we describe the identification of the first GPR30-specific agonist, G-1 (1), capable of activating GPR30 in a complex environment of classical and new estrogen receptors. The development of compounds specific to estrogen receptor family members provides the opportunity to increase our understanding of these receptors and their contribution to estrogen biology.

While acute myeloid leukemia (AML) comprises many disparate genetic subtypes, one shared hallmark is the arrest of leukemic myeloblasts at an immature and self-renewing stage of development. Therapies that overcome differentiation arrest represent a powerful treatment strategy. We leveraged the observation that the majority of AML, despite their genetically heterogeneity, share in the expression of HoxA9, a gene normally downregulated during myeloid differentiation. Using a conditional HoxA9 model system, we performed a high-throughput phenotypic screen and defined compounds that overcame differentiation blockade. Target identification led to the unanticipated discovery that inhibition of the enzyme dihydroorotate dehydrogenase (DHODH) enables myeloid differentiation in human and mouse AML models. In vivo, DHODH inhibitors reduced leukemic cell burden, decreased levels of leukemia-initiating cells, and improved survival. These data demonstrate the role of DHODH as a metabolic regulator of differentiation and point to its inhibition as a strategy for overcoming differentiation blockade in AML.