ALBERT

Description: Sickle cell disease (SCD) is an autosomal recessive genetic disorder caused by a point mutation in the human β-globin gene. Patients harboring this mutation can exhibit long-chain polymers of hemoglobin and sickle-shaped red blood cells, and suffer from severe medical manifestations including hemolysis and vaso-occlusive crises. Multiple preclinical, clinical and epidemiologic studies have shown that the levels of unmutated fetal hemoglobin (HbF encoded by the γ-globin gene) correlate with less severe disease, validating HbF induction as a therapeutic approach in SCD. Treatment with hydroxyurea (HU), the only approved therapy for SCD, results in a variable induction of HbF and significant improvement in the frequency of pain crises. However, a significant percentage of patients treated with HU fail to exhibit durable benefit, necessitating the need for alternative therapeutic agents. The human γ-globin gene is repressed in the post-natal period by epigenetic mechanisms, and therefore may lend itself to pharmacological intervention aimed at derepressing gene expression. One of the most important of these epigenetic mechanisms is catalyzed by lysine-specific demethylase 1 (LSD1), a histone demethylase that removes mono-/dimethyl marks from the lysine 4 and 9 residues of histone H3 through an FAD-directed redox process. Here, we report the characterization of selective, potent, and orally bioavailable LSD1 inhibitors from two classes - FAD-directed inhibitors that achieve inhibitory activity through formation of covalent FAD-adducts and non-FAD-directed, reversible inhibitors - and demonstrate their ability to induce γ-globin gene expression in murine and primate preclinical models. In the Towne's SCD mouse model, oral administration of LSD1 inhibitors significantly increased HbF+ cell (F cell) production. Concurrent with the increase in F cells, sickle cell numbers, reticulocyte counts, and bilirubin levels were all markedly reduced, indicating an amelioration of several pathophysiological features of SCD. FAD- and non-FAD-directed LSD1 inhibitors were more effective than HU in increasing F cells production, and the combination of HU and suboptimal doses of LSD1 inhibitors resulted in a greater induction of F cells and more pronounced reductions in reticulocyte counts and bilirubin levels. In addition to the humanized SCD model, HbF induction in response to LSD1 inhibitor treatment was evaluated in non-anemic cynomolgus monkeys. Oral administration of LSD1 inhibitors significantly induced F cells and HbF in a dose-dependent manner and over a sustained period (〉50 days) following the discontinuation of treatment. The percentage of induced F cells in total RBCs was linearly correlated with the percentage of HbF protein induced by LSD1 inhibition. Taken together, these results support the potential utility of LSD1 inhibition as a novel therapeutic approach to increase HbF production. Disclosures Lee: Incyte Corporation: Employment, Other: Stock. Soloviev:Incyte Corporation: Employment, Other: Stock. Zhang:Incyte Corporation: Employment, Other: Stock. Roman:Incyte Corporation: Employment, Other: Stock. Yang:Incyte Corporation: Employment, Other: Stock. Bowman:Incyte Corporation: Employment, Other: Stock. Burke:Incyte Corporation: Employment, Other: Stock. Margulis:Incyte Corporation: Employment, Other: Stock. O'Connor:Incyte Corporation: Employment, Other: Stock. Yang:Incyte Corporation: Employment, Other: Stock. Wu:Incyte Corporation: Employment, Other: Stock. Wynn:Incyte Corporation: Employment, Other: Stock. Burn:Incyte Corporation: Employment, Other: Stock. Shuey:Incyte Corporation: Employment, Other: stock. Diamond:Incyte Corporation: Employment, Other: Stock. Yao:Incyte Corporation: Employment, Other: Stock. Hollis:Incyte Corporation: Employment, Other: Stock. Yeleswaram:Incyte Corporation: Employment, Other: Stocks. Roberts:Incyte Corporation: Employment, Other: Stock. Huber:Incyte Corporation: Employment, Other: Stock. Scherle:Incyte Corporation: Employment, Other: Stock. Ruggeri:Incyte Corporation: Employment, Other: Stock.

Description: A significant population of patients with myelofibrosis (MF) develop anemia and either require red blood cell (RBC) transfusions or have an inadequate response to the currently available therapies and become transfusion-dependent. In patients with MF, elevated levels of serum hepcidin, a key iron regulatory hormone, is associated with increased dependence on RBC transfusions and reduced overall survival. Elevated hepcidin expression has also been observed to cause severe functional iron deficiency anemia and is central to the pathophysiology of anemia of chronic disease. Thus, to ensure proper maintenance of iron homeostasis, hepcidin levels are tightly regulated. Specifically, the production of hepcidin is controlled by the bone morphogenetic protein (BMP) type I receptor ACVR1, a gene that encodes the serine/threonine kinase ALK2. In preclinical models, knockdown or complete loss of ALK2 decreases hepcidin production resulting in elevated serum iron levels. In this study, we report characterization of INCB00928, a novel small molecule inhibitor of ALK2 for the treatment of anemia. INCB00928 was observed to have subnanomolar activity against ALK2 and selectivity over ALK1 and ALK3 in biochemical enzyme assays. In cell-based profiling studies, INCB00928 inhibited ALK2 potently and selectively over ALK1 and ALK3 as determined by the inhibition of ligand-induced SMAD pathway signaling. Importantly, in both an immortalized human liver cell line as well as primary human hepatocytes, INCB00928 inhibited BMP-induced production of hepcidin with nanomolar activity. INCB00928 was also observed to have suitable absorption, distribution, metabolism, and excretion properties to be dosed in in vivo rodent studies. In tumor- and inflammation-induced mouse models of anemia, INCB00928 improved RBC count, hemoglobin, and hematocrit levels while decreasing hepcidin levels in a dose-dependent manner. Additionally, consistent with the improved symptoms of anemia, pSMAD1/5 inhibition was observed in a dose-dependent manner in liver tissues collected from INCB00928-treated mice. In summary, INCB00928 is a potent, selective, and orally available small molecule inhibitor of ALK2, which significantly reduces the production of hepcidin in human liver cells, primary hepatocytes, and in rodent models of anemia. For the majority of patients with MF, the management of anemia remains an unmet need. The preclinical findings from this study suggest ALK2 kinase inhibition with INCB00928 may be a promising novel treatment to reduce the production of hepcidin and improve MF-related anemia in humans, thus warranting further investigation. Disclosures Chen: Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Stubbs:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Pusey:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Wen:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Collins:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Kapilashrami:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Rupar:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Thekkat:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Lin:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Bowman:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Yang:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Diamond:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Yeleswaram:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Kim:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Koblish:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Chen:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company. Wee:Incyte Corporation: Current Employment, Current equity holder in publicly-traded company.

Description: Indoleamine 2,3-dioxygenase-1 (IDO1; IDO) mediates oxidative cleavage of tryptophan, an amino acid essential for cell proliferation and survival. IDO1 inhibition is proposed to have therapeutic potential in immunodeficiency-associated abnormalities, including cancer. Here, we describe INCB024360, a novel IDO1 inhibitor, and investigate its roles in regulating various immune cells and therapeutic potential as an anticancer agent. In cellular assays, INCB024360 selectively inhibits human IDO1 with IC50 values of approximately 10nM, demonstrating little activity against other related enzymes such as IDO2 or tryptophan 2,3-dioxygenase (TDO). In coculture systems of human allogeneic lymphocytes with dendritic cells (DCs) or tumor cells, INCB024360 inhibition of IDO1 promotes T and natural killer (NK)–cell growth, increases IFN-γ production, and reduces conversion to regulatory T (Treg)–like cells. IDO1 induction triggers DC apoptosis, whereas INCB024360 reverses this and increases the number of CD86high DCs, potentially representing a novel mechanism by which IDO1 inhibition activates T cells. Furthermore, IDO1 regulation differs in DCs versus tumor cells. Consistent with its effects in vitro, administration of INCB024360 to tumor-bearing mice significantly inhibits tumor growth in a lymphocyte-dependent manner. Analysis of plasma kynurenine/tryptophan levels in patients with cancer affirms that the IDO pathway is activated in multiple tumor types. Collectively, the data suggest that selective inhibition of IDO1 may represent an attractive cancer therapeutic strategy via up-regulation of cellular immunity.