Abstract: The PI3K signaling pathway is frequently over-expressed in a variety of hematologic malignancies, so the development of PI3K inhibitors for the treatment of hematologic malignancies has broad application prospects. In this study, a novel PI3K inhibitor, JN-65, was identified through the investigation effects in the inhibition to hematologic malignancies. By MTT assays, JN-65 was found to effectively suppress the proliferation of hematologic malignancies, especially leukemia cell lines. The cell-free enzymatic studies demonstrated that JN-65 cloud inhibit PI3K and specifically inhibited PI3Kγ at low micromolar concentrations. Western blot confirmed that JN-65 could effectively inhibit the PI3K/Akt signaling pathway and the flow cytometry assays verified that JN-65 could induce the apoptosis of tumor cells through the suppression of PI3K signaling pathway. Finally, the molecular docking simulation method was used to explore the interaction between JN-65 and PI3K, and the inhibition mechanism of PI3K was revealed at the molecular level. In general, JN-65 would be a potential PI3K inhibitor for the treatment of hematologic malignancies.