Mechanisms of Resistance to the Fms-like Tyrosine Kinase 3 Inhibitor Crenolanib

Abstract

Mechanisms of Resistance to the Fms-like Tyrosine Kinase 3 Inhibitor Crenolanib: Trevor J. Mathias, Master of Science, 2014 Dissertation Directed by: Dr. Maria R. Baer, Professor, Department of Medicine and Molecular Medicine Acute myeloid leukemia (AML) is a bone marrow cancer in which myeloblasts fail to differentiate into downstream functional cell types. A number of genetic abnormalities can contribute to the development of AML. One of the most common and important is a mutation in the gene encoding Fms-like tyrosine kinase 3 (FLT3), a type 3 receptor tyrosine kinase, which is expressed on AML cells. The most common FLT3 mutation is an internal tandem duplication in the juxtamembrane domain, known as FLT3-ITD, resulting in constitutive and aberrant signaling. FLT3-ITD is present in ~30% of AML cases and is associated with poor treatment outcomes. Several FLT3 inhibitors are currently in clinical trials. One of these FLT3 inhibitors, crenolanib, is a potent and specific type I inhibitor, targeting the activated form of the kinase. My work is focused on the mechanisms of resistance and sensitization to crenolanib. Expression of ABC proteins associated with drug resistance on AML cells generally correlates with poor treatment response. I found that crenolanib is not an inhibitor of any of the three commonly upregulated ABC transporters, ABCB1, ABCG2, and ABCC1, but is a substrate of the ABCB1 transporter, indicating the potential for crenolanib to be effluxed from cells expressing ABCB1. Other FLT3 inhibitors have been shown to induce point mutations in FLT3-ITD, reducing the effectiveness of the treatment. Random mutagenesis in the presence of crenolanib produced point mutations in FLT3-ITD that only conferred mild resistance to crenolanib, with preserved sensitivity at effective and well tolerated concentrations in Phase I clinical trials. In cells containing FLT3-ITD there is increased expression of Pim-1, a proto-oncogene that is activated by a downstream target of FLT3, STAT5. Additionally, our laboratory has shown that Pim-1 promotes aberrant signaling of FLT3-ITD, and it is therefore an attractive therapeutic target. A Pim kinase inhibitor was used for in vitro combination treatments with crenolanib. This inhibitor synergized with crenolanib to produce increased levels of apoptosis. The mechanism behind this interaction is under further study. These studies helped to further elucidate the role of crenolanib in the treatment of FLT3-ITD positive AML.