Identification of AKN-032, a novel 2-aminopyrazine tyrosine kinase inhibitor, with significant preclinical activity in acute myeloid leukemia

• Published in: Biochemical pharmacology 2010-11, Vol.80 (10), p.1507-1516
• Main Authors: Eriksson, Anna, Höglund, Martin, Lindhagen, Elin, Åleskog, Anna, Hassan, Sadia Bashir, Ekholm, Carina, Fhölenhag, Karin, Jensen, Annika Jenmalm, Löthgren, Agneta, Scobie, Martin, Larsson, Rolf, Parrow, Vendela
• Format: Article
• Language: English
• Subjects: Acute myeloid leukemia; Adult; Aged; Animals; Antineoplastic Agents - adverse effects; Antineoplastic Agents - chemistry; Antineoplastic Agents - pharmacology; Antineoplastic Agents - therapeutic use; Apoptosis - drug effects; Biological and medical sciences; Cell Line, Tumor; Cell Survival - drug effects; Dose-Response Relationship, Drug; Epithelial Cells - drug effects; Epithelial Cells - enzymology; FARMACI; Female; Flow Cytometry; FLT3; fms-Like Tyrosine Kinase 3 - antagonists & inhibitors; Hematologic and hematopoietic diseases; Humans; Leukemia, Myelomonocytic, Acute - drug therapy; Leukemia, Myelomonocytic, Acute - enzymology; Leukemia, Myelomonocytic, Acute - pathology; Leukemias. Malignant lymphomas. Malignant reticulosis. Myelofibrosis; Male; Medical sciences; MEDICIN; MEDICINE; Mice; Middle Aged; Molecular Structure; New drug development; Pharmacology. Drug treatments; PHARMACY; Pyrazines - adverse effects; Pyrazines - chemistry; Pyrazines - pharmacology; Pyrazines - therapeutic use; Tyrosine kinase inhibitor; Xenograft Model Antitumor Assays; Young Adult
• ISSN: 0006-2952; 1356-1839; 1873-2968
• DOI: 10.1016/j.bcp.2010.08.002

Aberrant signal transduction by mutant or overexpressed protein kinases has emerged as a promising target for treatment of acute myeloid leukemia (AML). We here present a novel low molecular weight kinase inhibitor, AKN-032, targeting the FMS-like tyrosine kinase 3 (FLT3) and discovered in a new type of screening funnel combining the target therapy approach with sequential cellular screens. AKN-032 was identified among 150 selected hits from three different high throughput kinase screens. Further characterization showed inhibitory activity on FLT3 enzyme with an IC50 of 70nM. Western blot analysis revealed reduced autophosphorylation of the FLT3-receptor in AML cell line MV4-11 cells after exposure to AKN-032. Flow cytometry disclosed cytotoxic activity against MV4-11, but not against non-malignant 3T3-L1 fibroblast cells. Using a fluorometric microculture cytotoxicity assay, AKN-032 was tested against 15 cell lines and displayed a potent cytotoxic activity in AML cell lines MV4-11 (IC50=0.4μM) and Kasumi-1 (IC50=2.3μM). AKN-032 was also highly cytotoxic in tumor cells from AML patients in vitro. Furthermore, AKN-032 demonstrated significant antileukemic effect in a relatively resistant in vivo hollow fiber mouse model. No major toxicity was observed in the animals. In conclusion, AKN-032 is a promising new kinase inhibitor with significant in vivo and in vitro activity in AML. Results from the hollow fiber mouse assay suggest a favorable toxicity profile. Future studies will focus on pharmacokinetic properties, toxicity as well as further clarifying the mechanisms of action of AKN-032 in AML.