Novel thiazole amine class tyrosine kinase inhibitors induce apoptosis in human mast cells expressing D816V KIT mutation

Highlights

• D816V KIT expressing cells are sensitive to novel thiazole amine compounds.

• Synergy effect exists between thiazole amine compound and dasatinib and PKC412.

• The thiazole amine compounds inhibit β-catenin signaling.

Abstract

Gain-of-function mutations of receptor tyrosine kinase KIT play a critical role in the pathogenesis of systemic mastocytosis (SM) and gastrointestinal stromal tumors. D816V KIT mutation, found in ∼80% of SM, is resistant to the currently available tyrosine kinase inhibitors (TKIs) (e.g. imatinib mesylate). Therefore, development of promising TKIs for the treatment of D816V KIT mutation is still urgently needed. We synthesized thiazole amine compounds and chose one representative designated 126332 to investigate its effect on human mast cells expressing KIT mutations. We found 126332 inhibited the phosphorylation of KIT and its downstream signaling molecules Stat3 and Stat5. 126332 inhibited the proliferation of D816V KIT expressing cells. 126332 induced apoptosis and downregulated levels of Mcl-1 and survivin. Furthermore, 126332 inhibited the tyrosine phosphorylation of β-catenin, inhibited β-catenin-mediated transcription and DNA binding of TCF. Moreover, 126332 also exhibited in vivo antineoplastic activity against cells harboring D816V mutation. Our findings suggest thiazole amine compounds may be promising agents for the treatment of diseases caused by KIT mutation.

Introduction

Systemic mastocytosis is a group of heterogeneous disease characterized by abnormal proliferation and accumulation of mast cells in bone marrow, spleen, liver and gastrointestinal tract [1], [2]. Most of patients with indolent SM (ISM) respond well to the targeted therapy drugs and have a normal life expectancy [3]. Advanced SM includes aggressive SM (ASM), SM associated with clonal hematologic non-mast cell lineage disease (SM-AHNMD), and mast cell leukemia (MCL) according to the World Health Organization criteria [4], [5]. There are no effective therapies for patients with advanced SM. Approximately 80% of patients with advanced SM harbor the constitutively active D816V-mutated KIT, an acquired somatic mutation at codon 816 in KIT involving an aspartic acid to valine substitution [6].

KIT is transmembrane receptor tyrosine kinase. Under the physiological conditions, KIT is expressed in mast cells, stem cells, sperm cells and intestinal Cajal cells [7], [8]. KIT undergoes dimerization and autophosphorylation at tyrosine residues Y568 and Y570 of juxtamembrane domain becomes activated when binding of its ligand, stem cell factor [9]. This leads to the activation of signaling pathways such as Janus kinase/signal transducer and activator of transcription (JAK-Stat3/Stat5), mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K-Akt) [8]. Mutations in the KIT kinase domain changes the conformation of the KIT receptor, resulting in altered substrate recognition and constitutive tyrosine autophosphorylation, which leads to ligand-independent activation of KIT, leads to uncontrolled cell proliferation and a reduction in apoptosis [10], [11]. Gain of function mutations of KIT play critical role in the pathogenesis of mast cell leukemia (MCL), gastrointestinal stromal tumors (GISTs), and acute myelogenous leukemia (AML) [12].

KIT is an attractive therapeutic target because its oncogene addiction in patients with advanced SM and GISTs. Imatinib has been approved by U.S. Food and Drug Administration (FDA) for the treatment of adult SM patients without the KIT D816V mutation or with unknown mutational status [13], [14]. The results from clinical trials of dasatinib and nilotinib in patients with KIT D816V mutation were disappointing [15]. Midostaurin (PKC412) has synergistic effect with dasatinib and ponatinib against cells harboring the D816V mutation [16], [17]. Midostaurin exhibits promising efficacy for the treatment of ASM or MCL in a phase II clinical trial, however, the advantage of midostaurin over the current treatment such as interferon-α and cladribine is not clear. Novel TKIs such as semaxinib [18] and EXEL-0862 [19] have been reported to inhibit growth and induce apoptosis in mast cells harboring D816V KIT mutation, the long-term benefit for patients has not been determined. Therefore, development of promising novel TKIs for the treatment of aggressive SM and MCL is still urgently needed.

To overcome imatinib resistance, we have synthesized a series of thiazole amine compounds based on rationale design against imatinib-resistant KIT isoforms. Here, we evaluate the efficacy of two representative thiazole amine compounds designated 126332 and 126349. Our results showed that 126332 may be a promising agent for the treatment of diseases caused by KIT mutation.