Jason C. C. So

Bone marrow involvement by nasal NK cell lymphoma at diagnosis is uncommon.

To look for subtle evidence of marrow involvement in nasal NK cell lymphoma at diagnosis, we retrospectively studied trephine biopsy specimens from 25 consecutive patients by 2 sensitive techniques: CD56 immunohistochemistry and Epstein-Barr virus-encoded RNA in situ hybridization (EBER ISH). Only 2 patients had marrow involvement by NK cell lymphoma at diagnosis. In 3 additional patients, marrow involvement developed during or after systemic recurrence. All 5 positive cases were revealed by EBER ISH, but only 3 cases showed CD56 immunoreactivity. Among the 5 cases, only 2 were recognized by morphologic assessment. All 5 patients died, often within a short period, compared with a mortality of 50% for patients without demonstrable marrow involvement. Marrow involvement is distinctly uncommon in nasal NK cell lymphoma at diagnosis, and EBER ISH is the most sensitive technique for the demonstration of occult NK cell lymphoma. Despite the low frequency of marrow involvement in nasal NK cell lymphoma, EBER ISH is worthwhile to identify the minor subgroup of patients with a high likelihood of early death due to disease and when autologous bone marrow or peripheral blood stem cell transplantation is contemplated.

Mantle cell lymphoma in leukemic phase: characterization of its broad cytologic spectrum with emphasis on the importance of distinction from other chronic lymphoproliferative disorders.

Mantle cell lymphoma is a mature, virgin B‐cell neoplasm characterized immunologically by a panB+, CD5+, CD23−, cyclin D1+ phenotype and genetically by t(11;14)(q13;q32) with overexpression of the cyclin D1 (bcl‐1) gene. It usually presents as advanced stage disease, involving lymph nodes, spleen, bone marrow, and extranodal sites, particularly the gastrointestinal tract. However, frank leukemic presentation with high white cell counts is uncommon and can be difficult to distinguish from other chronic lymphoproliferative disorders. The aim of this study was to characterize the morphologic spectrum of leukemic mantle cell lymphoma.

Isolated extramedullary relapse after allogeneic bone marrow transplantation for chronic myeloid leukemia

Relapse of chronic myeloid leukemia (CML) as extramedullary granulocytic sarcoma (GS) after allogeneic bone marrow transplantation (BMT) is a rare occurrence. We report two patients who developed spinal GS as the first indication of relapse after allogeneic BMT for CML. In both cases, the marrow was in morphologic and karyotypic remission. However, fluorescence in situ hybridization (FISH) successfully demonstrated the presence of a minor Ph-positive clone in the marrow, as well as an occult clone with an additional Ph chromosome detected in one case. The results indicated a stronger graft-versus-leukemia effect in the marrow than in the peripheral tissues.

Indolent T-cell large granular lymphocyte leukaemia after haematopoietic SCT: a clinicopathologic and molecular analysis

Four women and three men after allogeneic (n=4) and autologous (n=3) haematopoietic SCT (HSCT) were observed to have an increase in T-cell large granular lymphocytes (T-LGLs) of CD3+CD8+ phenotype for a median of 41 (15–118) months. Clonal rearrangement of the T-cell receptor gene was verified by two PCR techniques and direct DNA sequencing, confirming that the cases were neoplastic and therefore classifiable as T-LGL leukaemia. In the allogeneic HSCT cases, T-LGL leukaemia was derived from donor T cells in three patients, as shown by DNA chimerism analysis, and recipient T cells in one patient who had graft failure previously. None of the patients showed cytopenia, autoimmune phenomenon or organ infiltration, which were features typical of de novo T-LGL leukaemia. Six patients had remained asymptomatic with stable large granular lymphocyte counts. One patient died from cerebral relapse of the original lymphoma. T-LGL leukaemias occurring post-HSCT are distinct from de novo T-LGL leukaemia and may have a different pathogenesis and clinical course. Patients did not require specific treatment, and the disease remained stable for long periods.

A unique case of B cell lymphoma relapsing as CD4/CD56 blastic neoplasm

A patient with history of B cell lymphoma treated with rituximab-based chemotherapy relapsed with a blastic CD4+/CD56+ neoplasm that was negative for CD20, CD79a and CD3. The relapse morphology and immunophenotyping were unusual and plasmacytoid dendritic cell (PDC) tumor enters the differential diagnosis. However, expressions of Oct-2 and CD10 in the relapse tumor were both more compatible with B cell than PDC lineage. Molecular investigations showed clonal rearrangements for both immunoglobulin heavy chain (IgH) and T cell receptor (TCR) γ chain gene by polymerase chain reaction (PCR). Furthermore, a clonal relationship with the original B cell lymphoma was demonstrated for all PCR products. Our case illustrated the potential pitfalls and ambiguity of lineage classification based on morphology and immunophenotyping alone, especially for rare and poorly defined entities.

Synchronous Epstein–Barr virus-positive diffuse large B-cell lymphoma of the elderly and Epstein–Barr virus-positive classical Hodgkin lymphoma

This work was partially funded by grants from the Ministerio de Sanidad y Consumo (PI05 ⁄ 1553, PI06 ⁄ 1074, and RD06 ⁄ 0020 ⁄ 0047), and Fundación MMA. P. Martı́n is in receipt of a research grant from the ISCIII. We thank Maria Lozano for her kind help with microdissection. The authors also wish to thank Martin Hadley-Adams for assisting with the English language and preparation of the manuscript.

Persistent neutropenia in chronic myelogenous leukemia in chronic phase treated with imatinib mesylate

A 48-year-old woman presented with left upper abdominal pain. Physical examination showed pallor and a 6-cm splenomegaly. The full blood count showed hemoglobin: 8.4 g/dL, white cell count: 144 x 10 9 /L, and platelet count: 982 x 10 9 /L. The blood smear showed a bimodal increase in myelocytes and neutrophils, with eosinophilia and basophilia. Bone marrow aspirate was markedly hypercellular with granulocytic hyperplasia and no increase in blasts. Megakaryocytes were small and hypolobated. Morphologic features were consistent with chronic myelogenous leukemia (CML) in chronic phase. However, cytogenetic analysis only showed 46, XX, inv(9)(p11q13).

Diagnosis of brainstem involvement in NK/T cell lymphoma

Dear Editor, A 71-year-old man was diagnosed in 1998 to have a nasal “T cell” lymphoma, which was treated with chemotherapy and radiotherapy. In March 2013, the lymphoma relapsed in his right orbit, with bilateral pulmonary infiltration shown on positron emission tomography–computed tomography (PET/ CT). A biopsy confirmed extranodal natural killer (NK)/T cell lymphoma, nasal type. He was initially treated with SMILE (dexamethasone, methotrexate, ifosfamide, L-asparaginase, etoposide) [1], but there was symptomatic progression after one course, and he was referred for further management. Salvage treatment with gemcitabine, cis-platinum, ifosfamide, L-asparaginase and etoposide was given. There was rapid relief of symptoms. After two courses of treatment, a complete response was demonstrated by PET/CT. He then declined further treatment. Two months later, he returned with multiple cranial nerve palsies on the left side. There was virtually complete ophthalmoplegia (Fig. 1a), left facial anaesthesia and deafness. Magnetic resonance imaging showed an enhancing lesion involving the left pons (Fig. 1b), cerebellar peduncle and temporal lobe. Enhancement and thickening of the left fifth cranial nerve was also found (Fig. 1c). The features were highly suggestive of lymphomatous involvement, but a biopsy of the involved sites was judged too risky neurologically. To reach a diagnosis, the cerebrospinal fluid (CSF) was examined. Predominantly small lymphocytes (47/μL) were found. However, very occasional large atypical lymphoid cells showing an irregular, nucleolated nucleus and an abundant amount of cytoplasm with azurophilic granules were observed (Fig. 1d, e). Although highly suspicious of being lymphomatous, these cells were too few for further characterization. To verify their neoplastic nature, quantitative polymerase chain reaction (Q-PCR) for Epstein-Barr virus (EBV) DNA calibrated with a World Health Organization EBV DNA standard [2] in the cell-free CSF was performed. The results showed EBV DNA at 5×10 IU/mL, providing confirmatory evidence of lymphoma involvement. However, a plasma sample taken at the same time did not show quantifiable EBV DNA. NK/T cell lymphomas are invariably infected by EBV [3]. When lymphoma cells undergo apoptosis, EBV DNA fragments are released. Hence, quantification of EBV DNA is a sensitive and accurate surrogate biomarker of lymphoma load, applicable to NK/Tcell lymphomas and other EBV-associated lymphoidmalignancies [2, 4]. EBVDNA of 10 to 10 IU/mL is typically found in the plasma of patients with disseminated NK/T cell lymphomas or aggressive NK cell leukaemia [2]. Hence, EBV DNA in the CSF of our patient was very high. Because there was minimal if any leptomeningeal involvement, the CSF EBV DNA was mainly derived from the brainstem lesions. Interestingly, the concomitant plasma EBVDNAwas negative, showing that the blood–brain barrier effectively prevented EBV DNA trafficking. This case illustrated two observations important in the management of NK/T cell lymphomas. Firstly, when lymphomatous involvement of a body cavity is suspected and cytological examination of the bodily fluid shows equivocal results, quantification of EBV DNA should be performed. Secondly, plasma EBV DNA quantification may not reflect lymphoma load in the central nervous system if the blood–brain barrier remains intact. J. C. C. So Department of Pathology, Queen Mary Hospital, Hong Kong, China

Unexpected glucose‐6‐phosphate dehydrogenase deficiency

A 50-year-old man with refractory intestinal T-cell lymphoma was started on combination chemotherapy (ifosfamide, methotrexate, l-asparaginase, etoposide, dexamethasone). Cotrimoxazole was commenced for pneumocystis prophylaxis after a normal glucose-6-phosphate dehydrogenase (G6PD) spot fluorescence screen [>4Æ5 iu/g haemoglobin (Hb)]. Initial tumour shrinkage was accompanied by renal impairment (creatinine 393 lmol/l, normal 67–109) and hyperuricaemia (1259 lmol/l, normal 260–530), and rasburicase (4Æ5 mg · 1 dose) was given for tumour lysis. Despite a dramatic fall in uric acid level (55 lmol/l), anaemia suddenly occurred (Hb fell from 113 g/l to 59 g/l), followed by hypotension and oliguria. Biochemical screening showed raised unconjugated bilirubin (41 lmol/l, normal <6) and lactate dehydrogenase (798 iu/l, normal 118–211). Intravascular haemolysis was confirmed by raised methaemalbumin (14Æ3 mg/l, normal <01Æ0) levels and haemoglobinuria. A peripheral blood film showed bite cells and abundant hemighost cells (left), classical for oxidative haemolysis. The patient survived with transfusion, haemodialysis and supportive treatment, with complete renal recovery. A repeat G6PD enzyme assay on his stored blood sample showed a marginal G6PD level of 5Æ5 iu/g Hb, which lies within the range expected for female heterozygotes. His four brothers were all G6PD deficient (0Æ21–1Æ24 iu/g Hb). Using allelespecific polymerase chain reaction, a G6PD Kaiping allele (nt 1388 G fi A) was detected in the patient and all siblings. Surprisingly the patient also carried a second normal G6PD allele, suggesting extra X chromosome material. On further questioning, the patient volunteered a history of azoospermia and bilateral absent vas deferens. There were no other abnormal features suggestive of Klinefelter syndrome. Karyotype analysis on stimulated peripheral blood lymphocytes and fluorescence in situ hybridization with an X whole chromosome painting probe (Oncor Inc., Gaithersburg, MD, USA) confirmed the presence of extra X chromosome material on the Y-chromosome (right, red arrow) in all 20 metaphases analysed: 46, XY.ish add(Y)(p11)(wcpX+). Hence the patient was an occult carrier of extra X chromosome material and a heterozygous carrier of G6PD deficiency. Deficiency of G6PD is the commonest red cell enzymopathy world-wide. It affects 4Æ8% of all Southern Chinese males, and is screened for at birth and before the commencement of oxidative medications. In female heterozygotes, random lyonization of the X chromosomes results in a mixture of normal and G6PD deficient red cells. The resultant average enzyme levels usually lie within the normal range. However female heterozygotes for G6PD deficiency can still suffer haemolysis as a result of skewed lyonization because of age, haemopoietic transplantation or clonal haemopoiesis. Without mandatory screening, this can cause unexpected severe haemolysis. Our case showed that a simple fluorescent screen might still be insufficient. Severe oxidative stresses because of a combination of drugs (cotrimoxazole and rasburicase) could still cause en masse destruction of the G6PD deficient subpopulation, resulting in life-threatening intravascular haemolysis. Hence, a quantitative enzyme level assay is a better option in areas endemic for G6PD deficiency.

A unique case of familial leukaemia: maternal puerperal leukaemia followed by infantile leukaemia with monosomy 7.

A 26-year-old mother (G1P1) presented with generalized lymphadenopathy 2 months after spontaneous vaginal delivery of a normal 3Æ5 kg daughter. A peripheral blood examination (haemoglobin 133 g/l, white cell count 4Æ2 · 10 /l, platelet count 183 · 10 /l) showed occasional blasts, and a marrow biopsy showed sheets of blasts (Fig 1A) that were negative for myeloperoxidase and non-specific esterase, but showed weak granular periodic acid-Schiff positivity. Flow cytometric immunophenotyping of the blasts showed expression of CD4, CD7, CD56 and HLA-DR, but negativity for Tdt, MPO, CD13, CD34 and other B and T cell markers (CD19, CD20, CD22 and CD2, CD3, CD5, CD8). Cytogenetics (Fig 1B) showed 44, XX, t(1;6)(q21;q23),)9,)13[5]/44,idem,t(9;17)(p22;p13)[3]/46XX[10]. She was treated as undifferentiated acute leukaemia with a lymphoid leukaemia protocol (Au et al, 1998) followed by allogeneic haemopoietic stem cell transplantation (HSCT) from an unrelated donor (two younger brothers were not a human leucocyte antigen (HLA) match) in first remission (conditioning: cyclophosphamide, total body irradiation). At 18 months of age, her infant daughter presented with recurrent fever and anaemia (haemoglobin 56 g/l, white cell count 5Æ5 · 10 /l, platelet count 393 · x10 /l). The peripheral film was unremarkable, but a marrow biopsy showed 14% granulated blasts and dysplastic megakaryocytes (Fig 1C), compatible with refractory anaemia with excess blasts, stage 2 (RAEB-2). Cytogenetic study showed 45, XX, )7[2]/46, XX[2]. Using a directly labelled chromosome 7 centromeric probe (D7Z1; Vysis, Downers Grove, IL, USA), monosomy 7 was shown in 58% of marrow nucleated cells (Fig 1D). Chromosome fragility screening using diepoxybutane-induced chromosome breakage was negative, but sequencing for RUNX1 mutation was not performed. The peripheral count was static for 6 weeks and the child underwent direct HSCT from another unrelated donor (conditioning: bulsuphan, cyclophasmide, melphalan, antithymocyte globulin). Both patients remained well at 2 and 1-year follow-up respectively. They denied any herbal or toxin exposure, nor any family history of leukaemia or malignancies; and neither patient had any dysmorphic features.