A Novel Axl And Mertk Dual Inhibitor Ono-7475: A New Therapeutic Agent For The Treatment Of Flt3-Itd And -Wild-Type Acute Myeloid Leukemia (Aml) Overexpressing

Introduction: The TAM (Tyro3, Axl, and Mertk) receptor tyrosine kinases (RTKs) together with their ligands Gas6 and Pros1 play diverse roles in survival signaling in cancers. Currently, the consensus indicates that therapeutic targeting of TAM RTKs will prove direct antitumor activity and may complement existing immunotherapy regimens to unleash the full power of the anticancer immune response. A number of small-molecule kinase inhibitors targeting TAM RTKs are being developed and tested in preclinical models. ONO-7475 is a highly potent and oral dual inhibitor of Axl and Mertk, currently in a phase 1 clinical study as a therapeutic agent for AML (NCT03176277). Our previous studies demonstrated that ONO-7475 induced complete tumor remission in the FLT3-internal tandem duplication (ITD)-positive AML cell line MV-4-11 mouse xenograft model (Yasuhiro et al., Blood 2014). However, the antitumor activity against FLT3-ITD AML cells was due to Axl inhibition (Ruvolo et al., Haematologica 2017) and the mechanism of Mertk regulation in AML has not been determined. To evaluate this question, we screened 28 AML-derived cell lines for FLT3-wild type (WT) and analyzed the influence of Mertk in proliferation. Methods: Twenty-eight FLT3-WT AML cell lines were treated with ONO-7475 for 72 hours, and the cell viabilities (IC50 values) were determined. Phospho-Axl (P-Axl), Axl and Mertk were determined by Western blot analysis. P-Mertk was determined by immunoprecipitation. Gene knockdown was achieved in FLT3-WT AML by transfecting with Axl, Mertk, Gas6, Pros1, or scrambled control siRNA. Results: Six out of 28 FLT3-WT cell lines responded to ONO-7475, and two cell lines, MOLM16 and MKPL1, were highly sensitive (IC50: 4.7nM and 83.5nM). Western blot analysis demonstrated that Axl, Mertk and P-Mertk were overexpressed in these ONO-7475-sensitive cells, and both are classified as megakaryoblastic leukemia (FAB-M7). SiRNA knockdown studies showed that the cell growth in MOLM16 was partially dependent on Mertk (63.5%) but totally dependent on double genes, Axl and Mertk. Furthermore, knockdown of the ligands Gas6 and Pros1 also partially inhibited the cell growth in MOLM16, respectively (46.0%, 49.2%). Conclusion: Our data suggest that Mertk may play an important role in survival signaling in FLT3-WT AML. Therefore, targeting both Axl and Mertk by ONO-7475 could be a new strategy for the treatment of FLT3-WT patients with AML in addition to FLT3-ITD. Additional work to investigate and clarify for target populations in both FLT3-ITD and -WT AML is currently under way. Citation Format: Kohei Tanaka, Cuifang Li, Tomomi Hirosaki, Hikaru Kato, Yoshinori Ishikawa, Miho Oka, Hiroshi Egawa, Ryohei Kozaki, Toshio Yoshizawa. A novel Axl and Mertk dual inhibitor ONO-7475: A new therapeutic agent for the treatment of FLT3-ITD and -wild-type acute myeloid leukemia (AML) overexpressing [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 1883.