Abstract 25: CG′806, a first-in-class FLT3/BTK inhibitor, exerts superior potency against AML cells harboring ITD, TKD and gatekeeper mutated FLT3 or wild-type FLT3

Abstract

Abstract The receptor tyrosine kinase FLT3 can undergo a series of mutations, including the activating internal tandem duplication (ITD) in the juxtamembrane region and point mutations in the tyrosine kinase domain such as at the activation loop residue D835 (Thiede et al. 2002). FLT3 is widely accepted as a prime target for acute myeloid leukemia (AML) therapy, as the FLT3-ITD mutation is present in approximately 24% of AML patients and it is associated with very poor prognosis (Kottaridis et al. 2003). However, additional acquired mutations of FLT3, including D835 or “gatekeeper” F691 mutations that have been identified in clinical patients who showed resistance/relapse to FLT3 inhibitors sorafenib or quizartinib (Man et al. 2012; Smith et al. 2012), can render most FLT3 inhibitors ineffective. We also reported that aberrant upregulation of other parallel prosurvival signaling pathways may render AML resistant to FLT3-targeted therapy (Zhang et al. 2014). CG′806 is a small-molecule multi-kinase inhibitor against FLT3 and BTK kinases that is under development to treat FLT3-driven AML. A single test concentration of 25 nM in a 583-kinase panel, an IC50 analysis against 176 kinases, and a Kd analysis against 483 kinases illustrated the ability of CG′806 to target the entirety of FLT3-mutant enzymes and to inhibit additional kinases (e.g., BTK, AURK, STE group, and TRK/AXL/DDR group). CG′806 exerted potent picomolar IC50 antiproliferative activity against human AML cells and against Ba/F3 mouse AML cells with FLT3-ITD mutations (about 50 to 250-fold higher activity compared to quizartinib or gilteritinib). Specifically, compared to second-generation FLT3 inhibitors quizartinib or gilteritinib, CG′806 showed much more pronounced antiproliferative effects in leukemia cells with D835 mutations, the ITD plus F691I/Y842D/D835 mutations, or in FLT3 wild-type cells (IC50s were 0.17, 0.82, 9.49, 0.30, 8.26, 9.72, and 0.43 nM for human ITD-mutated AML cells MV4-11 (FLT3-ITD), MOLM13 (FLT3-ITD), murine ITD mutated leukemia cells Ba/F3 WT, Ba/F3-ITD, Ba/F3-D835Y, Ba/FLT3-ITD+D835Y, and “gatekeeper” mutation Ba/F3-ITD+F691L cells, respectively). Furthermore, CG′806 triggered profound apoptosis in cell lines and primary AML patient samples harboring FLT3-ITD mutations as well as WT. Mechanistically, CG′806 profoundly suppressed FLT3 and its downstream MAPK/AKT signaling, as well as phospho-Aurora, and/or phospho-BTK proteins, suggesting the ability of CG′806 to inhibit various kinases that function in AML signaling and appear to contribute to its effectiveness. Moreover, CG′806 demonstrated in vivo tumor eradication without toxicity when administered orally, once daily for 14 d as a single agent in the MV4:11 AML murine xenograft model, and demonstrated sustained micromolar plasma drug levels in mice after a single oral administration. CG′806 can be considered not only a pan-FLT3 inhibitor for targeting FLT3 wild-type and ITD-mutant AML, but also a one-of-a-kind agent killing leukemia cells with TKD or dual TKD plus ITD mutations, which are frequently associated with resistance/relapse in FLT3-mutant AML patients. CG′806 exerts a robust therapeutic window in animal xenograft studies and does not appear to elicit the adverse side effects seen with other multikinase inhibitors (i.e., weight loss, nausea, diarrhea, myelosuppression, and/or vomiting). CG′806 warrants further investigation for the treatment of newly diagnosed and relapsed/refractory patients with FLT3-mutated AML. Citation Format: Weiguo Zhang, Hongying Zhang, Andrea Local, William G. Rice, Charlie J. Ly, Guopan Yu, Stephen B. Howell, Michael Andreeff. CG′806, a first-in-class FLT3/BTK inhibitor, exerts superior potency against AML cells harboring ITD, TKD and gatekeeper mutated FLT3 or wild-type FLT3 [abstract]. In: Proceedings of the Second AACR Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; May 6-9, 2017; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(24_Suppl):Abstract nr 25.