PS992 MSI2 REGULATES BACH1/BRIP1 THUS AFFECTING THE OXIDATIVE STRESS RESPONSE
Authors: J. Hoell, S. Duggimpudi, A. Borkhardt
Journal: HemaSphere (2019)

Background: AML is an aggressive hematopoietic malignancy that arises from a population of aberrant hematopoietic stem cells in the bone marrow (BM). Advances in understanding the molecular basis of AML has led to the development of new targeted therapies. CA-4948 is a novel, oral IRAK4 kinase inhibitor with additional inhibitory activity against wild-type (wt) and mutated FLT3 kinase. IRAK4 (Interleukin-1 Receptor Associated Kinase 4) is required for interleukin 1 receptor (IL-1R) and toll-like receptor (TLR) innate immune pathway signaling, pathways that are frequently over activated in AML and myelodysplastic syndromes (MDS). For example, AML patients have increased IL-1R agonist (IL-1ß) levels that promote the survival of AML cells and IL-1R KO represses AML cell growth in vitro and in vivo (Carey et al 2017). Dysregulation of the FLT3 signaling pathway is a well validated driver of AML. Constitutively activating mutations in FLT3 that comprise the ITD or the tyrosine kinase domain (KD) are frequently acquired late in AML disease and are poor prognostic factors with high relapse rates. FLT3 kinase inhibitors targeting FLT3-ITD or ITD/KD double mutations show high remission rates; however, multiple resistance mechanisms have been reported in both nonclinical models and AML patients. CA-4948 has both IRAK4 and FLT3 inhibitory activity, which may impart benefit to FLT3-wt and FLT3-mutant AML patients. Aims: Evaluate the ability of a novel IRAK4/FLT3 inhibitor, CA-4948, to block IRAK4 and FLT3 in FLT3-wt and FLT3-mutant AML in vitro and in vivo. Methods: CA-4948's kinome profile was assessed against 378 kinases and 9 FLT3 mutant variants (DiscoverX). CA-4948's ability to inhibit TLR/IL-1R or FLT3 signaling pathways was evaluated using NF-kB reporter, cytokine production, or western blot. The growth inhibitory and pro-apoptotic activity of CA-4948 was tested in vitro against AML cell lines by viability assay or flow cytometry. For AML FLT3-wt in vivo efficacy, THP-1 cells were tail-vein injected into mice and animal survival and degree of AML cell engraftment in BM were monitored in CA-4948, FLT3i, or vehicle treated mice. For AML FLT3 mutant in vivo efficacy, subcutaneous MV4–11 and MOLM-14 FLT3-ITD and MOLM-14 double FLT3-ITD/KD tumor models were treated with CA-4948 or other FLT3i. Results: CA-4948 exhibited 23 nM (Kd) binding affinity for IRAK4, >500-fold selectivity against IRAK1, and high binding affinity for FLT3-wt, -ITD, -ITD/D835 V, and -ITD/F691L at 8–31 nM (Kd). CA-4948 blocked cellular TLR and IL-1R-stimulated signaling from 150–220 nM (IC50). Consistent with CA-4948's additional FLT3 inhibitory activity, CA-4948 induced apoptosis and exhibited cytotoxic activity against AML FLT3 mutant lines in vitro at 58–200 nM (IC50). In FLT3-wt in vivo studies, CA-4948 blocked THP-1 bone marrow engraftment, while FLT3i quizartinib (Q) and midostaurin (M) had no inhibitory effect. In AML FLT3-ITD tumors, CA-4948 induced tumor regression equivalent to Q and M. CA-4948 also induced tumor regression in FLT3-ITD/D835Y tumors similar to M and showed greater efficacy in FLT3-ITD/F691L tumors as compared to Q. Summary/Conclusion: These results demonstrate that targeting the IL-1R/TLR signaling pathway with IRAK4 inhibitor CA-4849 may be an effective therapeutic strategy in FLT3-wt AML. Furthermore, CA-4948's additional FLT3 inhibitory activity may repress the emergence of FLT3-mutant AML clones. CA-4948's dual IRAK4/FLT3 inhibitory activity merits further clinical investigation in FLT3-wt and FLT3-mutant AML/MDS.