Yoshiharu Miyata

Extramedullary T-lymphoid blast crisis of an ETV6/ABL1-positive myeloproliferative neoplasm with t(9;12)(q34;p13) and t(7;14)(p13;q11.2)

Dear Editor, Chromosomal translocations involving ETV6 at 12p13 and ABL1 at 9q34 are very rare, but are very rare but recurrent aberrations in hematological malignancies including BCR / ABL1-negative chronic myeloid leukemia (CML), myeloproliferative neoplasm (MPN), and acute leukemia [1, 2]. The resultant ETV6/ABL1 fusion protein is thought to play a crucial role in leukemic transformation by the constitutive activation of tyrosine kinase [1]. Similar to BCR /ABL1-positive CML, it has been reported that several cases of ETV6 /ABL1 -positive CML/MPN presented with blast crisis (BC) in lymph nodes as well as bone marrow [3–7]. Here, we describe the first case of extramedullary T-lymphoid BC of ETV6 /ABL1 -positive MPN. A 31-year-old man was admitted because of generalized lymphadenopathy and leukocytosis. Peripheral blood values were hemoglobin 113 g/L, platelets 438×10/L, and leukocytes 50.7×10/L with 20 % myelocytes, 17 % metamyelocytes, 10 % band forms, 32 % segmented neutrophils, and 11 % eosinophils. Bone marrow was hypercellular with myeloid hyperplasia and prominent eosinophilia, and compatible with MPN (Fig. 1a–c). Pathological examination of inguinal lymph nodes was consistent with T-lymphoblastic lymphoma (T-LBL) (Fig. 1d). Immunohistochemistry showed that lymphoma cells were positive for CD3, CD1a, and TdT (Fig. 1e). The patient was treated by dasatinib with hyper-CVAD protocol followed by myeloablative allogeneic stem cell transplantation (SCT). However, he died of idiopathic pneumonia syndrome, 11 months after initial diagnosis. G banding and spectral karyotyping of bone marrow cells on admission showed 46,XY,der(9)t(9;12)(q34;p13), del(12)(p13)[1]/46,sl,t(7;14)(p13;q11.2)[18]/47,sdl1,+19[1] (Fig. 2a). The karyotype of lymph node cells was similar as fol lows: 46,XY,t (7 ;14)(p13;q11.2) ,der(9) t (9 ;12) (q34;p13),del(12)(p13)[4]. Fluorescence in situ hybridization (FISH) on metaphase spreads of bone marrow cells detected the ETV6 /ABL1 fusion signal on the der(9)t(9;12)(q34;p13) (Fig. 2b). FISH on interphase nuclei of lymph node cells also confirmed the ETV6 /ABL1 fusion signals (Fig. 2c). Reverse transcription-polymerase chain reaction (RTPCR) demonstrated two types of ETV6 /ABL1 fusion transcripts (types B and A) only in bone marrow cells of the patient (Fig. 2d) [2]. Southern blot analyses of lymph node cells showed rearrangement of TRD@ Jδ1, but germline configuration of IGH@ JH, TRB@ Cβ1, and TRG@ Jγ (data not shown). We have detected a rare translocation der(9)t(9;12) (q34;p13) resulting in an ETV6 /ABL1 fusion gene. We confirmed an expression of the fusion transcripts in K. Yamamoto (*) :K. Yakushijin :Y. Miyata :Y. Sanada : A. Okamura :H. Matsuoka :H. Minami Division of Medical Oncology/Hematology, Department of Medicine, Graduate School of Medicine, Kobe University, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 650-0017, Japan e-mail: kyamamo@med.kobe-u.ac.jp

CD56-positive diffuse large B-cell lymphoma: possible association with extranodal involvement and bcl-6 expression

Abstract Among B-cell non-Hodgkins lymphomas, neural cell adhesion molecule/CD56 expression is exceptional. In this study, seven cases of CD56-positive diffuse large B-cell lymphoma (DLBCL) are described. The frequency of CD56-positive DLBCL was 7% in our hospital. Four of seven (57·1%) cases expressed both CD10 and bcl-6 suggestive of a germinal center B-cell phenotype. Six of seven (85·7%) cases expressed bcl-6. Two cases expressed aberrant T cell-associated antigens, one each of CD7 and CD8. However, none of these seven cases showed CD5 expression. No significant difference was observed between CD56-positive and CD56-negative DLBCL in terms of the five international prognostic index risk factors. However, all seven cases had at least one extranodal involvement and showed a good response to initial treatment. The predominance of extranodal involvement in our series may be associated with the adhesion-related function of CD56. A high frequency of bcl-6 expression may be associated with a more favorable clinical course and prognosis.

NANOG Expression as a Responsive Biomarker during Treatment with Hedgehog Signal Inhibitor in Acute Myeloid Leukemia

Aberrant activation of the Hedgehog (Hh) signaling pathway is involved in the maintenance of leukemic stem cell (LSCs) populations. PF-0444913 (PF-913) is a novel inhibitor that selectively targets Smoothened (SMO), which regulates the Hh pathway. Treatment with PF-913 has shown promising results in an early phase study of acute myeloid leukemia (AML). However, a detailed mode of action for PF-913 and relevant biomarkers remain to be elucidated. In this study, we examined bone marrow samples derived from AML patients under PF-913 monotherapy. Gene set enrichment analysis (GSEA) revealed that PF-913 treatment affected the self-renewal signature and cell-cycle regulation associated with LSC-like properties. We then focused on the expression of a pluripotency factor, NANOG, because previous reports showed that a downstream effector in the Hh pathway, GLI, directly binds to the NANOG promoter and that the GLI-NANOG axis promotes stemness and growth in several cancers. In this study, we found that a change in NANOG transcripts was closely associated with GLI-target genes and NANOG transcripts can be a responsive biomarker during PF-913 therapy. Additionally, the treatment of AML with PF-913 holds promise, possibly through inducing quiescent leukemia stem cells toward cell cycling.

A novel TRB@/NOTCH1 fusion gene in T‐cell lymphoblastic lymphoma with t(7;9)(q34;q34)

Background: In T‐cell acute lymphoblastic leukemia/lymphoma (T‐ALL/LBL), activating mutations of NOTCH1 are observed in more than 50% of cases, whereas the t(7;9)(q34;q34) involving NOTCH1 at 9q34 and TRB@ at 7q34 is an extremely rare but recurrent translocation. Patient: A 41‐year‐old male with a large mediastinal mass, pleural effusion, and lymphadenopathy was diagnosed as having T‐LBL. Lymphoma cells were positive for CD4, CD8, CD2, CD3, CD5, CD7, CD10, and TdT. Results: G‐banding and spectral karyotyping of pleural effusion cells showed 47,XY,dup(1)(q21q32),t(7;9)(q34;q34),+20. Genomic polymerase chain reaction (PCR) revealed that the 5′ end of TRB@ J1‐5 was connected with the middle of NOTCH1 exon 25 (434 bp downstream from its 5′ end) in a ‘head‐to‐head’ configuration on the der(9)t(7;9), although nine extra bases were inserted between the two genes. Reverse transcription‐PCR confirmed expression of the TRB@/NOTCH1 fusion transcripts. Similarly, the 5′ end of J1‐5 was fused to the shortened exon 25 with nine extra bases. The NOTCH1 breakpoint in exon 25 was very close to transcription start sites of deleted Notch1 in murine T‐ALL. Conclusions: The TRB@/NOTCH1 fusion gene with a NOTCH1 breakpoint in exon 25, which has not previously been detected in four other reported cases with t(7;9), could lead to aberrant expression of the truncated NOTCH1 by TRB@ enhancer elements. The resultant NOTCH1 receptor deleting most of the extracellular domain may be implicated in the pathogenesis of T‐LBL by ligand‐independent, constitutive activation of the NOTCH1 pathway, suggesting avenues for future therapy with γ‐secretase inhibitors.

An mTORC1/2 kinase inhibitor enhances the cytotoxicity of gemtuzumab ozogamicin by activation of lysosomal function

Gemtuzumab ozogamicin (GO), the first antibody-drug conjugate (ADC), has attracted the interest of hematologists because more than 90% of acute myeloid leukemia (AML) blasts express its target, CD33. Although GO and subsequently developed ADCs depend on lysosomes for activation, lysosome number and activity in tumor cells has not been well elucidated. In this study, we investigated whether an mTORC1/2 kinase inhibitor, PP242, which was reported to activate lysosomal function, potentiates the cytotoxicity of GO in AML cells. Eight AML cell lines (U937, THP-1, SKM-1, SKK-1, SKNO-1, HL-60, MARIMO and KO52) were treated with GO and PP242. The cytotoxic effect of GO was enhanced by concurrent treatment with a non-cytotoxic concentration (500 nM) of PP242 in most cell lines, except MARIMO and KO52 cells. We then used LysoTracker to label acidic lysosomes in U937, THP-1, SKM-1, MARIMO and KO52 cells. LysoTracker fluorescence was dramatically increased by treatment with PP242 in U937, THP-1 and SKM-1 cells, and the intensified fluorescence was retained with PP242 + GO. In contrast, PP242 did not induce a significant increase in fluorescence in MARIMO cells, consistent with the lack of combinatory cytotoxicity. LysoTracker fluorescence was also increased by PP242 in KO52 cells, which have been reported to strongly express multidrug resistance (MDR). Further, PP242 suppressed GO-induced Chk1 activation and G2/M cell cycle arrest, which in turn triggered cell cycle promotion and cell death. These results indicate that inhibition of mTORC1/2 kinase by PP242 enhanced the cytotoxicity of GO by increasing lysosomal compartments and promoting the cell cycle via suppression of GO-induced Chk1 activation. This combination may represent an attractive new therapeutic strategy for the treatment of leukemia.

A Case of Classical Hodgkin Lymphoma with Total Lymph Node Infarction

Lymph node infarction is very rare, and is frequently associated with neoplasms, such as malignant lymphoma and non-neoplastic disease, or interventions such as fine-needle aspiration (FNA). A 76-year-old-man presented with cervical lymph node swelling. Although FNA was performed, the findings were insufficient for a definitive diagnosis. Consequently, surgical biopsy of the cervical lymph node was performed, which revealed total infarction; a diagnosis of classical Hodgkin lymphoma was made later. Both lymphoma itself and FNA may cause total lymph node infarction, which makes diagnosis confusing. Therefore, it is important to repeat the biopsy rather than repeat FNA to correctly diagnose malignant lymphoma, including Hodgkin lymphoma.

Invasive Scopulariopsis alboflavescens infection in patient with acute myeloid leukemia

Scopulariopsis alboflavescens is a soil saprophyte that is widely distributed in nature. Recently, there have been increasing number of reports of invasive infections with Scopulariopsis species in immunocompromised patients. In this report, we described an adult woman with acute myeloid leukemia and who developed S. alboflavescens pneumonia. Liposomal amphotericin B and voriconazole combination therapy was unsuccessful and the patient died because of pneumonia. Scopulariopsis is highly resistant to available antifungal agents and almost invariably fatal. This case report should alert clinicians to the importance of listing Scopulariopsis as a pathogenic fungus in immunocompromised patients.

Unbalanced Translocation der(7)t(7q;11q): A New Recurrent Aberration Leading to Partial Monosomy 7q and Trisomy 11q in Acute Myeloid Leukemia

munophenotypically positive for CD13, CD33, CD34 and HLA-DR. Considering dysplastic changes of bone marrow cells, we made a diagnosis of AML with myelodysplasia-related changes (MRC). The patient received an induction therapy with aclarubicin and low-dose cytarabine followed by six courses of azacitidine therapy, but he could not achieve complete remission. He died of progressive disease 10 months after the initial diagnosis. G-banding and spectral karyotyping of bone marrow cells at diagnosis showed 46,XY,der(7)t(7; 11)(q11.2;q13) [18]/46,sl,add(3)(p11)[2] ( fig. 1 a, b). The karyotype after 6 months was also 46,XY,der(7)t(7; 11)(q11.2;q13)[20]. Fluorescence in situ hybridization (FISH) on metaphase spreads detected three MLL signals on the der(7)t(7; 11) and two normal chromosomes 11 ( fig. 1 c), whereas one D7S486 signal at 7q31 was deleted from the der(7)t(7; 11) ( fig. 1 d). These findings confirmed that the der(7)t(7; 11) (q11.2;q13) as a sole abnormality resulted in partial monosomy 7q and partial trisomy 11q including MLL amplification without rearrangement [5, 6] . In addition, we examined the possible involvement of RUNX1 at 21q22 by FISH on interphase nuclei, because De Braekeleer et al. [5] revealed an abnormal clone with a cryptic RUNX1 deletion, which was distinct from the clone with der(7)t(7; 11)(q31;q14), in AML at diagnosis. However, Unbalanced translocations, which are created if one of the two derivative chromosomes is lost, often result in the loss or gain of chromosomal material rather than the formation of fusion genes in hematological malignancies [1] . They are usually part of complex karyotypes, diagnostically nonspecific and related to disease progression, although some of them seem to be a recurrent and primary anomaly [2, 3] . Recently, unbalanced translocations involving chromosome 11, such as der(18)t(11; 18) (q13;q21.1) and der(7)t(7; 11)(q31;q14), have been reported in acute myeloid leukemia (AML) [4, 5] . These translocations occurred as a sole abnormality at diagnosis, and resulted in partial trisomy 11q including three copies of the MLL gene at 11q23. Here, we describe a new case of AML with der(7)t(7q;11q), which led to partial monosomy 7q and partial trisomy 11q including MLL . A 79-year-old Vietnamese male was admitted because of anemia. He had no history of chemoradiotherapy. Peripheral blood showed Hb 7.0 g/dl, PLT 44 × 10 9 /l and WBC 18.5 × 10 9 /l with 25% myeloblasts, 6% myelocytes, 16% metamyelocytes, 5% band forms, 24% segmented neutrophils, 1% basophils, 12% monocytes and 11% lymphocytes. Bone marrow was hypercellular with 22.1% myeloblasts, 66.2% myeloid cells and 0.8% erythroblasts. Blasts were positive for myeloperoxidase staining and imReceived: December 9, 2013 Accepted: December 20, 2013 Published online: May 22, 2014