Rock Y. Y. Leung

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.

Non-nasal natural killer cell lymphoma: not non-nasal after all.

Dear Editor, Natural killer (NK) cell lymphoma, or extranodal NK/T-cell lymphoma, nasal type, according to the World Health Organization classification, is a rare neoplasm affecting preferentially Asian and South American populations, being extremely unusual in Western patients [1]. There are two clinical forms, nasal and non-nasal (extranasal) [2]. In nasal NK-cell lymphoma, the nasal and upper aerodigestive areas are initially involved. Dissemination to the skin, gut, testis, salivary glands or marrow occurs in advanced diseases. In non-nasal NK-cell lymphomas, however, the skin, gut, testis, salivary glands or marrow are the primary sites. Non-nasal NK-cell lymphomas have been reported to have a worse prognosis [3]. A 59-year-old man presented with bone pain, jaundice and fever. There were no lymphadenopathy or organomegaly. Investigations showed hemoglobin: 11.4 g/dL, white cell count: 1.4×10/L with no abnormal circulating cells, platelet count: 45×10/L, bilirubin: 67 (7–19) μmol/L, alkaline phosphatase: 510 (49–138) U/L, alanine aminotransferase: 407 (6–53) U/L, aspartate aminotransferase: 336 (13–33) U/L and lactate dehydrogenase (LDH): 1,200 (107– 218) U/L. A positron emission tomography/computerized tomography (PET/CT) showed hyper-metabolic lesions confined exclusively to the bone marrow (Fig. 1a). Histologic examination of the marrow showed diffuse infiltration by medium-sized lymphoid cells with irregular nuclei, clumped chromatin and cytoplasm with fine azurophilic granules, associated with florid hemophagocytosis (Fig. 1b). The neoplastic cells were surface CD3–, CD2+, CD7+, CD56+, and strongly positive for Epstein Barr virus encoded RNA on in situ hybridization (Fig. 1c). A nasal panendoscopy showed normal nasal, nasopharyngeal and oropharyngeal areas. Overall features were consistent with non-nasal NK-cell lymphoma. He was treated with methotrexate 2 g/m (day 1), and ifosamide 1.5 g/m/day, etoposide 100 mg/m/day, dexamethasone 40 mg/day (days 2 to 4) [4]. He responded with normalization of temperature and LDH. On day 8, however, he developed fever, nasal pain and obstruction with epistaxis. A CT scan showed a 2-cm rim-enhancing irregular lesion in the nasopharynx, a site which was uninvolved initially (Fig. 1d). Biopsy of the lesion confirmed NK-cell lymphoma infiltration (Fig. 1e). Treatment with Lasparaginase (6,000U/m/alternate daily×7) induced complete resolution of symptoms. Following normalization of blood counts and liver function, a marrow examination showed complete morphologic remission. Nasal panendoscopy with random nasopharyngeal biopsies also showed normal findings. Although NK-cell lymphomas are considered aggressive [1], stage I/II nasal NK-cell lymphomas have a favorable prognosis with radiotherapy and chemotherapy [5], whereas stage III/IV nasal NK-cell lymphomas fare much worse with few survivors. Conversely, non-nasal NK-cell lymAnn Hematol (2009) 88:185–187 DOI 10.1007/s00277-008-0562-0

Invasive fungal infections after obinutuzumab monotherapy for refractory chronic lymphocytic leukemia

Dear Editor, Patient 1 was a 40-year-old man with refractory chronic lymphocytic leukemia (CLL), having received in the past 8 years R-CHOP, FCR, R-B, DHAP, BEAM, MINE (supplemental file), alemtuzumab, and radiotherapy. He was persistently neutropenic and had three septicemic episodes responding to broad-spectrum antibiotics. In January 2014, the anti-CD20 antibody obinutuzumab was administered via a named patient program (NPP) together with antifungal (itraconazole) and anti-pneumocytis (cotrimoxazole) prophylaxis. After the third dose of obinutuzumab, fever not responding to broad-spectrum antibiotics and echinocandin (micafungin) developed. Blood cultures were negative. Later, chest X-ray and computed tomographic scan showed diffuse ground-glass consolidations (Fig. 1a). Bronchoalveolar lavage yielded Pneumocystis jirovecii. Although high-dose cotrimoxazole was administered, fever persisted. A repeat blood culture showed Candida krusei. Despite high-dose liposomal amphotericin B, he died of multi-organ failure. Patient 2 was a 38-year-old man with refractory small lymphocytic lymphoma/CLL, having received in the past 6 years FC (supplemental file), FCR, R-CHOP, and R-B. He was persistently neutropenic, but no septic episodes had occurred. In February 2014, obinutuzumabwas administered via an NPP, with inhalational pentamidine prophylaxis. After the third dose of obinutuzumab, fever not responding to broad-spectrum antibiotics and echinocandin (anidulafungin) developed. Blood cultures were negative. Bone marrow examination showed septated fungi (Fig. 1b), confirmed by culture to be Penicillium marneffei. High-dose liposomal amphotericin B and itraconazole were administered. Fever failed to respond. His chest X-ray then showed bilateral shadowing (Fig. 1c), associated with progressive shortness of breath. It was highly suspicious of P. jirovecii infection, but severe thrombocytopenia precluded bronchoscopic confirmation. High-dose cotrimoxazole was given, resulting in resolution of fever and X-ray changes. A fourth course of obinutuzumab was administered under cotrimoxazole and itraconazole coverage. Before obinutuzumab treatment, both patients had been heavily pre-treated with potent lymphodepleting/cytotoxic drugs, resulting in prolonged leucopenia. However, with appropriate prophylaxis, none of them developed opportunistic infections. Yet, once obinutuzumab was given, both patients developed invasive fungal infections (IFI). In case 1, despite prophylactic cotrimoxazole, P. jirovecii infection still occurred. Furthermore, candidemia developed despite the use of itraconazole and micafungin. In case 2, an even more serious disseminated P. marneffei infection developed. P. marneffei infection is an acquired immunodeficiency syndrome (AIDS) defining condition [1]. Similar to case 1, despite prophylactic pentamidine, an infection highly likely due to P. jirovecii developed. The apparent failure of prophylaxis might be related to severe lymphopenia. In patient 1, the leucocyte counts were between 0.2 and 0.4×10/L, practically all of which were abnormal lymphoid cells. In patient 2, the CD4 Tcell count at diagnosis of P. marneffeiwas 0.04×10/L. Hence, in both cases, the CD4 T cell counts would be lower than 0.05×10/L, which defined severe lymphopenia as occurring in AIDS. IFI is rarely attributed to the type I anti-CD20 antibody rituximab [2]. For the type II anti-CD20 antibody obinutuzumab, trials in newly diagnosed CLL [3] and relapsed/refractory B cell lymphomas [4, 5] had not reported any IFI. Obinutuzumab is much more potent than rituximab in E. Tse :Y.<L. Kwong (*) Department of Medicine, Queen Mary Hospital, Pokfulam Road, Hong Kong, China e-mail: ylkwong@hkucc.hku.hk

CD20 expression in natural killer T cell lymphoma

Zhaoli Lane Robert William Allan Peng Wang Charles Chuanhai Guo Yan Peng Jianping Li Department of Pathology and Immunology, Washington University School of Medicine, St Louis, Department of Pathology, Henry Ford Hospital, Detroit, MI, Department of Pathology, Immunology and Laboratory Medicine, Gainesville, FL, USA, Department of Pathology, Beijing Ditan Hospital, Beijing, China,Department of Pathology, University of Texas MD Anderson Cancer Center, Houston, and Southwestern Medical Center, University of Texas, Dallas, TX, USA

Isolated gingival relapse of acute lymphoblastic leukemia after transplantation.

A patient with a history of precursor B-cell leukemia presented with an isolated ulcerating gum lesion 8 years after allogeneic stem cell transplantation with severe graft versus host disease. A biopsy revealed an undifferentiated malignant hemic lesion. Examinations of the peripheral blood and marrow were normal. Molecular studies confirmed clonal relationship between the gum lesion with the original marrow disease, despite the anatomical, histological and chronological separations.

Diffuse Osteosclerosis Complicating Hairy Cell Leukemia

A 67-year-old man presented with mild pancytopenia 3 years ago, for which no specific treatment was given. Progressive anemia necessitated a bone marrow examination 2 years later. Aspiration yielded a dry tap, and a trephine biopsy showed marked osteosclerosis with myelofibrosis. On referral, physical examination was normal. Blood counts showed hemoglobin of 8.3 g/dL, WBC count of 2.8 10/L (neutrophils, 1.8 10/L; lymphocytes, 0.9 10/L; monocytes, 0.08 10/L), and platelet count of 81 10/L. No abnormal cells were discernable on the blood film. Dual-energy x-ray absorptiometry showed massively increased bone mass density (BMD), particularly over the axial skeleton and long bones (Fig 1A, arrows). The measured BMDs at different regions were as follows: lumbar spine, 2.28 g/cm (11 standard deviations [SDs] above the mean for young adults); femur, 1.95 g/cm (eight SDs above mean); and total body, 1.61 g/cm (5.6 SDs above mean; Fig 1B). A positron emission tomography– computed tomography confirmed the osteosclerosis, but no solid tumors were detectable. Bone marrow biopsy was repeated. Severe osteosclerosis (Fig 2A, arrows, hematoxylin and eosin staining) was present, along with a dense infiltrate of abnormal lymphoid cells, showing a typical spaced appearance, with pale to clear areas surrounding the nuclei (Fig 2A, insert, red arrows). Severe reticulin fibrosis (Fig 2B, silver staining) was also present. Immunohistochemical analysis showed that the lymphoid cells were positive for CD20 and acid phosphatase isoenzyme 5 (tartrate resistant; Fig 2C, immunoperoxidase staining). A review of the peripheral-blood buffy coat showed rare abnormal lymphoid cells with hairy projections (Fig 2D, arrow, Wright-Giemsa staining). The overall findings were consistent with hairy cell leukemia (HCL). Serum level of the osteoclastogenesis inhibitory factor osteoprotegerin (assayed by enzyme-linked immunosorbent assay; Biomedica Gruppe, Vienna, Austria) was grossly elevated at 11.5 pmol/L (normal adult male, 0.55 to 2.35 pmol/L). The patient received one course of subcutaneous 2-chlorodeoxyadenosine (0.14 mg/kg/d 5). Blood counts gradually normalized. A marrow trephine biopsy 4 months later showed persistence of osteosclerosis, but no abnormal lymphoid cells could be found. The BMD had remained unchanged (total body, 1.68 g/cm), which was an expected result because bone turnover typically takes much longer. Except for unusual cases of late-onset osteopetrosis, diffuse osteosclerosis in adults is almost invariably associated with underlying neoplasms. Metastases from prostate and breast cancer and advanced primary myelofibrosis are important causes. Osteosclerosis

Macrophage activation syndrome leading to fatality in subcutaneous panniculitis-like T cell lymphoma

Dear Editor, Subcutaneous panniculitis-like T cell lymphoma (SPTCL) is rare. It is generally indolent with favourable treatment results. About 20 % of cases are complicated by a haemophagocytic syndrome (HPS), which portends a poor outcome. However, reasons for the poor survivals in SPTCL with HPS are undefined. A 19-year-old man presented with multiple painless nodules over his body. Positron emission tomography–computed tomography (PET-CT) showed widespread abdominal lesions and multiple lymphadenopathy. A skin biopsy showed SPTCL. He was treated with prednisolone, retinoids and interferon alpha. One year afterwards, progressive abdominal distension, night sweating and weight loss occurred. A CT scan showed hepatosplenomegaly. Bone marrow examination was interpreted as showing lymphomatous infiltration. He was treated with one course of CHOP (cyclophosphamide, doxorubicin, vincristine, prednisolone) without response and was referred. On referral, there were pancytopenia, deranged liver function, hypofibrinogenaemia, elevated lactate dehydrogenase and extreme hyperferritinaemia (287,600 pmol/L; reference range,

Unremitting pyrexia, pancytopenia, hepatosplenomegaly, and extreme hyperferritinemia

A 52-year-old woman presented with unremitting pyrexia reaching 40 C. There was progressive weight loss and jaundice. A series of investigations failed to give a diagnosis, and because of rapid deterioration she was referred for management. On admission, she was moribund, with a fever of 40 C. There was marked pallor and jaundice. Abdominal examination showed gross hepatomegaly and splenomegaly (22 cm and 6 cm below the costal margin, respectively). A full blood count showed hemoglobin: 5.5 g/dL, white cell count: 1.42 3 10/L, and platelet count: 19 3 10/L. Liver function showed bilirubin: 47 mmol/L (reference range: 4–23 mmol/L), alanine aminotransferase (ALT): 68 IU/L (7–36 IU/L), alkaline phosphatase (ALP): 1,088 IU/L (32–93 IU/L), and albumin: 24 g/L (39–50 g/L). The lactate dehydrogenase (LDH) was 785 IU/L (107–218 IU/L). The renal function test was normal. Coagulation tests showed prothombin time (PT): 19.1 sec, activated partial thromboplastin time (APTT): 63.2 sec, fibrinogen: 0.29 g/L (1.46–3.38 g/ L), and D-dimer: 31.04 mg/L (<0.95 mg/L). The pancytopenia prompted a bone marrow examination. The marrow aspirate showed marked lymphohistiocytic infiltration and frequent hemophagocytosis (Fig. 1A). The constellation of unexplained pyrexia, hepatosplenomegaly, pancytopenia, hypofibrinogenemia, and marrow hemophagocytosis was consistent with the syndrome of hemophagocytic lymphohistiocytosis (HLH). HLH is a hyperinflammatory condition caused by unchecked and dysregulated stimulation of the immune system [1–3]. HLH is classified etiologically as primary and secondary (Table I). Primary HLH pertains to patients who have a history of genetic diseases, and is caused by gene mutations leading to defects in proteins that play important roles in the cytolytic secretory pathway in natural killer (NK) and cytotoxic Tcells (CTL). These defects result in impaired elimination and control of intracellular pathogens, leading to prolonged and enhanced stimulation of the immune system. NK-cells and CTLs secrete large amounts of cytokines that, besides causing a systemic inflammatory response, also activate macrophages, resulting in their infiltration into various organs and hemophagocytosis. Primary HLH manifests in infancy and early childhood, often due to a triggering infectious agent such as Epstein–Barr virus (EBV). Rarely, presentation in adulthood may occur [4]. Secondary HLH is a group of heterogeneous disorders, which is reactive to underlying infections, malignancies, and systemic autoimmune diseases. EBV and tuberculosis are common causes of infection-related HLH [2]. Lymphomas, predominantly T-cell or NK-cell lymphomas, are the most common underlying malignancies [2]. The diagnosis of HLH is based on the HLH-2004 criteria, which stipulate a combination of five or more of the following manifestations: pyrexia; splenomegaly; cytopenia affecting two or more lineages in the peripheral blood; hypertriglyceridemia and/or hypofibrinogenemia; hemophagocytosis in bone marrow, spleen, or lymph nodes; low or absent NK-cell activity; hyperferritinemia, and increased soluble CD25 [2]. Alternatively, molecular features consistent with HLH can be used to make the diagnosis. The presentation of our patient fulfilled these diagnostic criteria. The advanced age of our patient implied that the HLH was secondary. She was investigated for secondary causes of HLH. Autoimmune serologic markers were normal. A computed tomography (CT) showed gross hepatosplenomegaly with multiple enlarged para-aortic lymph nodes (Fig. 2A). Microbiological and serological tests for infections were negative. The serum ferritin level was markedly elevated to 179,500 pmol/L, which was about 250 times the upper reference limit (reference range: 52–738 pmol/L). Ferritin is an acute phase reactant, and hyperferritinemia is found in various inflammatory and neoplastic conditions. The degree of hyperferritinemia provides diagnostic clues (Table II). Mild infections, inflammatory conditions, liver diseases and autoimmune disorders rarely result in ferritin of >5 times normal [5]. Ferritin of higher levels might be found in patients with malignancies, particularly lymphoproliferative diseases. Much higher levels of ferritin are found in HLH and lymphomas [1– 3]. Macrophages are the main source of serum ferritin [6]. In HLH, macrophage activation by massive release of cytokines, including interleukin-1 (IL-1), IL-6, and tumor necrosis factor alpha (TNF-a), also leads to secretion of ferritin from macrophages, accounting for the hyperferritinemia. Few conditions result in very high (>50 times) levels of ferritin, and iron overload and severe infections such as disseminated tuberculosis [7] should be considered. For extreme hyperferritinemia (>100 times), juvenile idiopathic arthritis and adult onset Still’s disease (AOSD) are virtually the only known causes [8]. Extreme hyperferritinemia rendered AOSD a likely diagnosis. A profile of serum cytokines was obtained,

Post-rituximab Burkitt transformation of PTLD: loss of CD20 expression accompanied by a switch in light-chain expression.

Dear Editor, A 53-year-old woman, with cadaveric liver transplant for primary biliary cirrhosis 7 years ago, presented with generalized lymphadenopathy (Fig. 1a). A biopsy showed post-transplantation lymphoproliferative disease (PTLD; Fig. 1b). The lymphoma cells expressed CD20, CD79a, κ-chain, and Epstein Barr virus-encoded RNA (EBER). Her bone marrow examination was normal but a serum immunoelectrophoresis showed a monoclonal protein (IgGλ 8.2 g/l). Cyclosporin was stopped, and she was treated with R-CEOP×6 (rituximab, cyclophosphamide, epirubicin, vincristine, and prednisolone). This was followed by acute liver rejection, requiring tacrolimus, prednisolone, and mycophenolate mofetil treatment. A remission lasted for 4 years, with undetectable circulating EBV-DNA and monoclonal protein. However, she relapsed with fever, renal shutdown, and hepatosplenomegaly (Fig. 1c). A marrow aspirate showed sheets of Burkitt-like cells (Fig. 1d), expressing CD19, CD22, CD23, EBER, and λ-chain, but not CD10, CD20, CD34, and FMC-7. There was spontaneous tumor lysis (lactate dehydrogenase 4,760 IU/ml, normal 200–360; urate 1,120 mmol/l, normal 160–380), accompanied by an upsurge in circulating EBVDNA and a new monoclonal band (IgMκ 4.6 g/l). She was treated with COMP×4 (cyclophosphamide, vincristine, methotrexate, and prednisolone). Tacrolimus was continued but she died of isolated brain relapse 6 months later. The two EBV-related lesions had different morphology, immunophenotype, light-chain expression, and clinical behavior. Interestingly, in both instances, the PTLD lightchain expression was discordant with the concomitant circulating M-band, although the presence of monoclonal proteins and PTLD are known to be unrelated after most solid transplants [1]. The clinical history is best explained by continued selection among multiple subclones of abnormal EBV-driven B cells since initial presentation. Polymerase chain reaction (PCR) analysis was performed on DNA extracted from the presenting and relapsing lesions (Fig. 1e) using concensus sets of immunoglobulin heavy chain primers covering the three hypervariable complementary determining regions and three conserved framework regions [2]. The existence of different subclones was confirmed by different PCR product sizes in the two samples in two experiments. Since the VH3 PCR products showed two bands for each sample, to resolve the issue of biclonal disease versus bi-allelic arrangement, the products were cloned, sequenced, and aligned as reported (Fig. 1e and f) [3]. All four bands were clonally related (Fig. 1g, left), supporting biclonal evolution within each sample, and divergent evolution from a common occult EBV immortalized ancestral B-cell clone (Fig. 1g, right) [4]. From the clinical point of view, the initial dramatic and sustained clinical response of the disseminated PTLD Ann Hematol (2010) 89:97–99 DOI 10.1007/s00277-009-0769-8

Pyrexia, Massive Hepatomegaly, and Extreme Hyperferritinemia

CLINICAL SUMMARY Investigations showed a hemoglobin level of 9.3 g/dL, leukocyte level of 2.3 10/L, platelet level of 162 10/L, bilirubin level of 76 (4-23) mmol/L, alanine aminotransferase level of 94 (7-36) IU/mL, aspartate aminotransferase level of 94 (14-30) IU/mL, and alkaline phosphatase level of 824 (32-93) IU/mL. Serologic markers for autoimmune diseases were negative. Serum ferritin, measured by a 2-site chemiluminometric assay with the World Health Organization standard 80/578, was elevated to 883,100 (25-275) pmol/L. Bone marrow examination showed hemophagocytosis (Figure 1A, B) and 10% atypical lymphoid cells. Polymerase chain reaction for T-cell receptor alpha, beta, and gamma genes showed polyclonal bands, excluding a T-cell lymphoma. Positron emission tomography and computed tomography showed eumetabolic massive hepatomegaly (Figure 1C, D). Liver biopsy showed moderate mixed macrovesicular and microvesicular steatosis (Figure 1E, F). Two months after presentation, hepatic failure developed, and liver allografting was considered. However, there was improvement after prednisolone (1 mg/kg) therapy. One month later, a salmon-pink rash developed on the patient’s face and trunk. Adult-onset Still’s disease was diagnosed. She was treated with intravenous tocilizumab (8 mg/kg), resulting in defervescence within 1 day, liver decreasing to 10 cm within 1 week, and ferritin decreasing to 42,505 pmol/L. Her blood counts normalized after the second dose of tocilizumab. The addition of cyclosporin with tocilizumab (8 mg/kg every 4 weeks) ultimately led to complete