br Conflict of interest br Acknowledgments The project has b

Conflict of interest
Acknowledgments The project has been supported by grants from Science Foundation Ireland, Ireland (Starting Investigator Research Grant to ES, 09/SIRG/B1575, TIDA to ES 12/TIDA/B2388) and by the Millennium Fund of NUIG.
The t(3;21)(q26;q22) is a rare cytogenetic abnormality, reported in approximately 1% of all myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) cases and mainly occurring in therapy-related myeloid neoplasms, in accelerated or blast phase of chronic myeloid leukemia or other myeloproliferative neoplasms (MPN), and, rarely, in AML . Previous treatment with hydroxycarbamide or other antimetabolites is strongly implicated as a contributory cause . The presence of t(3;21)(q26;q22), isolated or associated with other chromosomal abnormalities, is associated with a very poor prognosis . In this translocation, portions of the RUNX1 gene have been reported to be variably fused to MECOM (currently preferred designation for the dextromethorphan hydrobromide located within the 3q26 region, namely RPL22, MDS1, EVI1), as a result of alternative intergenic splicing, with production of multiple fusion transcripts . RUNX1–MECOM (formerly AML1–MDS1–EVI1) fusion products directly contribute to leukemogenesis or leukemic transformation, can block myeloid differentiation and promote proliferation by exerting a dominant-negative effect over RUNX1-induced normal transcriptional activation, antagonize the growth-inhibitory effects of transforming growth factors, block JNK activity and therefore prevent stress-induced apoptosis, and enhance AP-1 activity . We report here the unusual case of an elderly patient observed because of persisting splenomegaly and increased lactate dehydrogenase (LDH), heralding AML with eosinophilia and RUNX1–MECOM fusion transcripts. In details, a 63-year old Caucasian man with previous history of minor β-thalassemia, arterial hypertension, hypothyroidism, fibromyalgia, benign prostatic hyperplasia and acute myocardial infarction in 2009 was admitted to our outpatient׳s Hematology Unit in April 2013 because of splenomegaly and persistently increased LDH, mainly LDH2 isoform, since 2009. The blood examinations documented white blood cell (WBC) count 4.7×10/L, with a differential count showing 47% neutrophils, 34% lymphocytes, 9% monocytes, 5% eosinophils, 5% basophils, hemoglobin (Hb) level 15.4g/dl with MCV 76fl, platelet (Plt) count 338×10/L, serum LDH 1053IU/L. The morphological examination of the peripheral blood (PB) smear did not document cytological abnormalities, while neutrophil alkaline phosphatase activity was elevated. Neither abnormal myeloid cells nor atypical lymphocytes were detected by flow cytometry on PB samples. Microbiological, virological and autoimmunity examinations were negative. Homogeneous splenomegaly (bipolar diameter 16.2cm) without signs of portal hypertension was observed on ultrasonography. Neither bone marrow (BM) examination nor cytogenetic and molecular analyses were performed at this time-point, whereas the morphological examination of a BM aspirate performed in 2008 because of transient mild neutropenia was unrevealing. In August 2013 the patient was subsequently admitted because of fever, fatigue, abdominal pain and drenching night sweats. The laboratory investigations revealed WBC count 12.2×10/L with a differential count showing 25% neutrophils, 11% lymphocytes, 2% monocytes, 43% eosinophils, 1% basophils and 18% blasts, Hb level 9.7g/dl, Plt count 17×10/L, LDH 548IU/L. Unfortunately, serum tryptase level was not measured. Homogeneous splenomegaly was worsened (bipolar diameter 20cm), as observed on abdominal ultrasonography. The morphological examination of BM aspirate (A) and trephine biopsy (B) showed hypercellularity (90%), with marked proliferation of eosinophil granulocytopoiesis (40%), multilineage myelodysplastic features, especially with small hypolobated megakaryocytes, and a blast cell count 20–30%. Blast cells were minimally differentiated, with high nuclear/cytoplasmic ratio and basophilic cytoplasm. They were CD34+, CD33+/−, CD13+, CD117+, HLA-DR+, CD38+, c-MPO− by flow cytometry analysis performed on the BM aspirate. Moreover, mild BM fibrosis was documented on Gomori methenamine silver staining (C). The presence of dense mast cell collections or atypical spindle-shaped mast cells was morphologically excluded. Moreover, neither immunohistochemical examinations for CD68 and CD117 nor flow-cytometry analyses to investigate the expression of CD2, CD25 and CD117, performed on BM trephine biopsy and BM aspirate, respectively, documented atypical or aggregated mast cells (not shown). Unfortunately, immunostaining for tryptase was not performed. Based upon these features, AML with MDS-related changes and eosinophilia was thus diagnosed. Conventional G-banding showed 46,XY,t(3;21)(q26;q22) karyotype in all the 20 metaphase cells analyzed (D). Molecular examinations, namely reverse transcriptase (RT)-PCR (E) and subsequent sequencing analyses (F) performed on both PB and BM samples, also documented RUNX1–MECOM, alternatively spliced multiple fusion transcripts .