The FIP1L1-PDGFRA fusion gene and the KIT D816V mutation are coexisting in a small subset of myeloid/lymphoid neoplasms with eosinophilia

To the editor:

Spindle-shaped tryptase⁺/CD25⁺/CD117⁺/CD2^+/− mast cells (MCs) are a hallmark of the bone marrow (BM) in myeloid/lymphoid neoplasms harboring a FIP1L1-PDGFRA (FP) fusion gene.^1,2  These neoplasms that may present as chronic eosinophilic leukemia (CEL), acute myeloid leukemia/blast phase, myeloid sarcoma, or lymphoblastic lymphoma/leukemia rarely fulfill the criteria of the World Health Organization category “systemic mastocytosis with associated haematological non-mast cell disorders” (SM-AHNMD).^1-5  Among 123 SM-AHNMD patients, only 12 carried an FP fusion gene.⁵ KIT D816V analysis performed on 8 of these FP-positive patients and on 2 cases included in another study were negative.^5,6  Thus, the relationship between MCs in FP-positive CEL and KIT D816V-positive SM remains elusive.

We microdissected tryptase⁺ MCs from formalin-fixed, EDTA-decalcified, paraffin-embedded BM trephine biopsies of 19 FP-positive CEL patients using a PALM MicroBeam (Carl Zeiss Microscopy, Oberkochen, Germany). DNA was extracted from 100 to 250 pooled MCs per biopsy (QIAamp DNA Mini Kit; Qiagen, Hilden, Germany). KIT D816V mutation was evaluated by analyzing KIT exon 17 nested polymerase chain reaction products by LightCycler^-based high-resolution melting and conventional direct sequencing.⁶  Genomic DNA from the KIT D816V-positive MC leukemia cell line HMC-1 and NB4 cells harboring wild-type KIT served as controls. The polymerase chain reaction reactions and final results were evaluated in a blinded fashion. Low levels of KIT D816V-mutated DNA ranging from <10% to 20% were detected in the MCs of 5 of 19 FP-positive CEL patients (Figure 1). The KIT D816V mutation was not detectable in DNA/RNA extracted from whole white blood cells derived from peripheral blood or BM samples, probably as a consequence of the very low burden of mutated KIT D816V-positive cells, which was only detectable in DNA obtained from microdissected MCs. Fluorescence Immunophenotyping and Interphase Cytogenetic as a Tool for Investigation Of Neoplasia (FICTION) using a Vysis 4q12 tri-color rearrangement fluorescence in situ hybridization probe kit (Abbott Molecular, Wiesbaden, Germany) resulted in 1 green/aqua fusion signal with a deletion of the orange signal of the CHIC2 gene in the nuclei of tryptase⁺ BM MCs in a patient with KIT D816V-positive MCs (Figure 1). The presence of CD25⁺-positive MCs with concomitant KIT D816V and serum tryptase levels >20 ng/mL support the diagnosis of SM-AHNMD/SM-FP–positive CEL in 2 patients. This discrepancy to the previously published negative KIT D816V mutational analysis of microdissected MCs may be explained by a sampling effect because only 2 FP-positive SM-CEL cases were examined.⁶  It remains elusive whether FP and KIT D816V are present in the same clone or whether there are 2 separate clones. We reported on the heterogeneity of molecular aberrations in KIT D816V-positive SM with ≥1 additional mutation, for example, TET2, SRSF2, ASXL1, and others in 24 of 27 patients.⁷  In a murine model, expansion of eosinophils and MCs may result from an interaction between FP, interleukin 5, the ligand stem cell factor, and KIT in the absence of a KIT mutation.⁸  It can be speculated that the FP fusion gene favors secondary KIT mutations in MCs via growth and proliferation signals or that a yet unknown mechanism causes genomic instability with independent evolution of FP and KIT D816V.