Abstract Acute myeloid leukemia (AML) is the most prevalent adult leukemia characterized by genetic or mutational disruption of myeloid differentiation, growth arrest, and apoptosis. Standard “7 + 3” chemotherapy consisting of cytarabine in combination with an anthracycline such as daunorubicin has been used for more than four decades. However this course only results in 10 year disease free survival in 15% of patients age <60 years and 2% among those > 60 years. Several new successful therapies in AML are shown to work by promoting differentiation, and/or inducing apoptosis of the terminally differentiated cells. However these therapies, while promising, have the limitation of working in only a subset of AML with specific genetic profiles. The discovery of novel agents applicable to a broader patient population still represents a critical unmet need in this disease. Intracellular calcium is a common signaling molecule used in a variety of cellular processes including cell cycle progression. One of the major pathways which regulates calcium entry is store operated calcium entry (SOCE) mediated through calcium release-activated Ca2+ channels (CRAC), which respond to depleted calcium stores in ER lumen. This primarily occurs via interaction of ORAI1 (expressed on the plasma membrane) and STIM1 (expressed on the endoplasmic reticulum). Leukemic cells in particular often rely on the influx of calcium through CRAC for proliferation, therefore targeting these channels are a promising therapeutic option. In the current study, we explore the preclinical use of RP4010, a novel inhibitor of ORAI1 developed by Rhizen Pharmaceuticals, in AML. RP4010 is very effective at inhibiting proliferation of both AML cell lines and primary AML patient samples (IC50 values ranging from 0.2 - 1 μM using a tetrazolium-based colorimetric assay (MTS) assay). Importantly, RP4010 efficacy is not altered in the presence of stromal cell support. RP4010 also decreased colony growth in CFU assays. Finally, we evaluated the in vivo efficacy using an MV4-11 xenograft model. RP4010 treatment at 50 mg/kg once daily by oral gavage significantly prolonged survival compared to the vehicle control (p = 0.019). Furthermore, at the time of sacrifice, splenic tumor burden (percentage of human CD45 positive cells) was reduced in the mice treated with RP4010 compared to the vehicle group (p = 0.019). In conclusion, we provide here compelling evidence that inhibition of ORAI1 with RP4010 is a promising therapeutic strategy in AML. A Phase 1/1b study in R/R Non-Hodgkin Lymphoma (NHL) patients is currently underway in the US (ClinicalTrials.gov Identifier: NCT03119467), and our data would support expansion to other hematological malignancies as well, particularly AML. Senior authors J. C. B and E. H contributed equally to this work. Citation Format: Bridget Carmichael, Amy Lehman, Ola A. Elgamal, Shelley J. Orwick, Jean Truxall, Larry Beaver, Kumar V. Penmetsa, Srikant Viswanadha, John C. Byrd, Erin Hertlein. Store-operated calcium signaling is an effective therapeutic target in AML [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1938.