A novel K509I mutation of KIT identified in familial mastocytosis—in vitro and in vivo responsiveness to imatinib therapy
Introduction
KIT is the cellular homologue of v-kit, which is found in the Hardy-Zuckerman 4 feline sarcoma virus [1] and is a Class III transmembrane receptor tyrosine kinase (RTK). Following ligand binding to the Ig domains, dimerisation occurs thus activating intrinsic kinase activity by transphosphorylation of specific residues in the JM and TK domains. KIT mutation has been implicated in a wide range of tumour types and although many KIT mutations have been identified to date, they appear to cluster in only a few sites in the molecule.
Exon 11 mutations affect the juxtamembrane (JM) domain, in particular the tyrosine residues 550–560 that form an inhibitory alpha helix, which suppresses autophosphorylation. Mutations of this region disrupt the binding of regulatory proteins to the JM region, result in autophosphorylation and constitutive activation of the tyrosine kinase. Such activating mutations have been seen in mast cell lines [2], [3] as well as small numbers of adults with mast cell disease [4]. Gastrointestinal stromal tumours (GISTs) are also commonly associated with exon 11 mutations.
Exon 17 mutations affect the TK domain with the most commonly observed mutation being an A > T substitution at nt 2648 resulting in a D816V substitution. This stabilises the kinase loop in its active configuration and hence induces activation of STAT3 and STAT1 and up-regulates downstream targets of STAT3 such as Bcl-xL and c-myc [5]. The mast cell line HMC-1 contains the D816V mutation resulting in ligand-independent activation and cellular transformation [3]. TK mutations are frequent in cases with mast cell expansion and the incidence of KIT mutation is higher in patients with systemic mastocytosis compared to those with cutaneous mastocytosis.
The expanding variety of identified KIT mutations is of importance with the advent of novel tyrosine kinase inhibitors. Imatinib is a tyrosine kinase inhibitor that targets Bcr-Abl, PDGFR-α and -β and Kit. Imatinib inhibits wild type Kit and JM mutants but has no effect on D816V [6] due to conformational changes of the drug's binding site. Imatinib has been used in 12 cases of mast cell disease [7] resulting in 5 complete remissions. Notably the failures were in the two patients with the D816V mutation. A proportion of cases of mastocytosis with eosinophilia are now known to have the FIP1L1-PDGFRA fusion that is imatinib sensitive [8]. Imatinib sensitivity has additionally been recently demonstrated in a case of sporadic mastocytosis bearing a novel germline F522C transmembrane mutation of exon 10 [9]. This is at difference to the previous cases discussed. In this paper we report a case of familial mastocytosis where in vitro and in vivo sensitivity to imatinib was demonstrated. Subsequent genetic study demonstrated the absence of the D816V mutation, consistent with imatinib responsiveness, and further identified a novel K509I mutation in exon 9 in both of the affected patients.
Section snippets
Patients
Two related patients are currently under the care of the Department of Haemato-Oncology at St. Bartholomew's Hospital with aggressive and indolent mastocytosis. The family tree is demonstrated in Fig. 1.
Patient II2 presented to our department in July 1981, aged 26, with hepatosplenomegaly. She had previously been well, apart from an undiagnosed childhood skin complaint, an appendicectomy in 1968 and an axillary vein thrombosis in 1971. Examination of her skin revealed dermatographism and
Imatinib sensitivity testing
Sensitivity testing to imatinib for patient II2 revealed that in liquid culture 1 μM induced no change in viability as compared to that of the control; however, 5 μM resulted in a reduction in viability from day 12 onwards (Fig. 2(a)). For semisolid culture, reduction in colony counts for both CFU-GM and CFU-Baso was noted in the presence of imatinib for the BM-MNC from patient II2 as compared to control MNC cells from healthy individuals (Fig. 2(b)).
Clinical response to imatinib
The results of in vitro work suggested some
Discussion
Familial mastocytosis is a very rare condition with perhaps only 50 families recorded in the literature [15], [16]. It is usually inherited in an autosomal dominant manner, presenting with cutaneous disease in infancy or childhood that may then persist. Variable systemic involvement is noted. Discordance has been noted for monozygotic twins so additional factors may be necessary [17].
In this study we have identified a K509I change in the juxta-membrane domain of KIT present in the affected
Acknowledgements
NC and LYZ were supported by the Leukemia Research Fund and the Wessex Cancer Trust, JVM and BS were supported by the Leukemia Research Fund and MS, JF and TAL were supported by Cancer Research UK.
Contributions. LYZ and NC provided detailed mutational screening of KIT and developed the ARMS assay; BS and JVM provided in vitro imatinib sensitivity data; MS, JF, TAL and JDC contributed to the clinical care of the patients and their initial molecular evaluation; MS wrote paper and coordinated
References (25)
- et al.
Constitutive activation of c-kit in FMA3 murine mastocytoma cells caused by deletion of seven amino acids at the juxtamembrane domain
Blood
(1996) - et al.
Identification of activating c-kit mutations in adult-, but not in childhood-onset indolent mastocytosis: a possible explanation for divergent clinical behavior
J Invest Dermatol
(1998) - et al.
Signal transducer and activator of transcription 3 activation is required for Asp(816) mutant c-Kit-mediated cytokine-independent survival and proliferation in human leukemia cells
Blood
(2001) - et al.
FIP1L1-PDGFRA fusion: prevalence and clinicopathologic correlates in 89 consecutive patients with moderate to severe eosinophilia
Blood
(2004) - et al.
Diagnostic criteria and classification of mastocytosis: a consensus proposal
Leuk Res
(2001) - et al.
Src family kinases are involved in the differential signaling from two splice forms of c-Kit
J Biol Chem
(2003) - et al.
Achalasia due to diffuse esophageal leiomyomatosis and inherited as an autosomal dominant disorder. Report of a family study
Gastroenterology
(1990) - et al.
Tyrosine kinase oncogenes in normal hematopoiesis and hematological disease
Oncogene
(2002) - et al.
Identification of mutations in the coding sequence of the proto-oncogene c-kit in a human mast cell leukemia cell line causing ligand-independent activation of c-kit product
J Clin Invest
(1993) - et al.
Effect of tyrosine kinase inhibitor STI571 on the kinase activity of wild-type and various mutated c-kit receptors found in mast cell neoplasms
Oncogene
(2003)
Imatinib for systemic mast-cell disease
Lancet
A novel form of mastocytosis associated with a transmembrane c-Kit mutation and response to imatinib
Blood
Cited by (125)
Drug-induced mast cell eradication: A novel approach to treat mast cell activation disorders?
2022, Journal of Allergy and Clinical ImmunologyCitation Excerpt :Therefore, imatinib may be considered as an MC-depleting therapy in such patients, especially in those with a well-differentiated MC morphology. Indeed, major responses or even remissions have been reported in these patients during imatinib therapy.82-86 It should also be noted that imatinib is a US Food and Drug Administration– and European Medicines Agency–approved drug for this indication (KIT D816V–negative SM).
Clinical impact and proposed application of molecular markers, genetic variants, and cytogenetic analysis in mast cell neoplasms: Status 2022
2022, Journal of Allergy and Clinical ImmunologyDeveloping a standardized approach for assessing mast cells and eosinophils on tissue biopsies: A Work Group Report of the AAAAI Allergic Skin Diseases Committee
2021, Journal of Allergy and Clinical ImmunologyPathogenic and diagnostic relevance of KIT in primary mast cell activation disorders
2021, Annals of Allergy, Asthma and ImmunologyGenome-wide association study identifies novel susceptibility loci for KIT D816V positive mastocytosis
2021, American Journal of Human Genetics