Pruritic arthropod bite-like papules in T-cell large granular lymphocytic leukaemia and chronic myelomonocytic leukaemia.

T-cell large granular lymphocytic leukaemia (T-LGLL) is a clinically indolent mature T-cell neoplasm characterized by a monoclonal population of CD3+ CD8+ cytotoxic T cells, which usually presents as neutropenia, anaemia and thrombocytopenia. Chronic myelomonocytic leukaemia (CMML) is a clonal haematopoietic disorder with features of both a myeloproliferative neoplasm and myelodysplastic syndrome (MDS). Patients with CMML exhibit a persistent peripheral blood monocytosis in addition to myelodysplastic features. Because of the rarity of T-LGLL, its cutaneous manifestations are poorly documented, but include vasculitis, vasculopathy, persistent ulcerations, generalized pruritus and disseminated granuloma annulare. Various types of skin lesions have been observed in patients with CMML and reportedly occur in approximately 10% of cases. We report the extraordinary case of a patient with MDS who developed T-LGLL, and subsequently the MDS progressed to CMML. The patient then developed diffuse arthropod bite-like papules and intractable pruritus.

Regulation of PI3K signaling in T-cell acute lymphoblastic leukemia: a novel PTEN/Ikaros/miR-26b mechanism reveals a critical targetable role for PIK3CD.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy, and T-ALL patients are prone to early disease relapse and suffer from poor outcomes. The PTEN, PI3K/AKT and Notch pathways are frequently altered in T-ALL. PTEN is a tumor suppressor that inactivates the PI3K pathway. We profiled miRNAs in Pten-deficient mouse T-ALL and identified miR-26b as a potentially dysregulated gene. We validated decreased expression levels of miR-26b in mouse and human T-ALL cells. In addition, expression of exogenous miR-26b reduced proliferation and promoted apoptosis of T-ALL cells in vitro, and hindered progression of T-ALL in vivo. Furthermore, miR-26b inhibited the PI3K/AKT pathway by directly targeting PIK3CD, the gene encoding PI3Kδ, in human T-ALL cell lines. ShRNA for PIK3CD and CAL-101, a PIK3CD inhibitor, reduced the growth and increased apoptosis of T-ALL cells. Finally, we showed that PTEN induced miR-26b expression by regulating the differential expression of Ikaros isoforms that are transcriptional regulators of miR-26b. These results suggest that miR-26b functions as a tumor suppressor in the development of T-ALL. Further characterization of targets and regulators of miR-26b may be promising for the development of novel therapies.

Loss of PRDM1/BLIMP-1 function contributes to poor prognosis of activated B-cell-like diffuse large B-cell lymphoma.

PRDM1/BLIMP-1, a master regulator of plasma-cell differentiation, is frequently inactivated in activated B-cell-like (ABC) diffuse large B-cell lymphoma (DLBCL) patients. Little is known about its genetic aberrations and relevant clinical implications. A large series of patients with de novo DLBCL was effectively evaluated for PRDM1/BLIMP-1 deletion, mutation, and protein expression. BLIMP-1 expression was frequently associated with the ABC phenotype and plasmablastic morphologic subtype of DLBCL, yet 63% of the ABC-DLBCL patients were negative for BLIMP-1 protein expression. In these patients, loss of BLIMP-1 was associated with Myc overexpression and decreased expression of p53 pathway molecules. In addition, homozygous PRDM1 deletions and PRDM1 mutations within exons 1 and 2, which encode for domains crucial for transcriptional repression, were found to show a poor prognostic impact in patients with ABC-DLBCL but not in those with germinal center B-cell-like DLBCL (GCB-DLBCL). Gene expression profiling revealed that loss of PRDM1/BLIMP-1 expression correlated with a decreased plasma-cell differentiation signature and upregulation of genes involved in B-cell receptor signaling and tumor-cell proliferation. In conclusion, these results provide novel clinical and biological insight into the tumor-suppressive role of PRDM1/BLIMP-1 in ABC-DLBCL patients and suggest that loss of PRDM1/BLIMP-1 function contributes to the overall poor prognosis of ABC-DLBCL patients.

Differential PAX5 levels promote malignant B-cell infiltration, progression and drug resistance, and predict a poor prognosis in MCL patients independent of CCND1.

Reduced Paired box 5 (PAX5) levels have important roles in the pathogenesis of human B-cell acute lymphoblastic leukemia. However, the role of PAX5 in human lymphoma remains unclear. We generated PAX5-silenced cells using mantle cell lymphoma (MCL) as a model system. These PAX5(-) MCL cells exhibited unexpected phenotypes, including increased proliferation in vitro, enhanced tumor infiltration in vivo, robust adhesion to the bone marrow stromal cells and increased retention of quiescent stem-like cells. These phenotypes were attributed to alterations in the expression of genes including p53 and Rb, and to phosphoinositide 3-kinase/mammalian target of rapamycin and phosphorylated signal transducer and activator of transcription 3 pathway hyperactivation. On PAX5 silencing, the MCL cells displayed upregulated interleukin (IL)-6 expression and increased responses to paracrine IL-6. Moreover, decreased PAX5 levels in CD19+ MCL cells correlated with their increased infiltration and progression; thus, PAX5 levels can be used as a prognostic marker independent of cyclin D1 in advanced MCL patients. Importantly, high-throughput screening of 3800 chemical compounds revealed that PAX5(-) MCL cells are highly drug-resistant compared with PAX5 wild-type MCL cells. Collectively, the results of our study support a paradigm shift regarding the functions of PAX5 in human B-cell cancer and encourage future efforts to design effective therapies against MCL.

Comprehensive gene expression profiling and immunohistochemical studies support application of immunophenotypic algorithm for molecular subtype classification in diffuse large B-cell lymphoma: a report from the International DLBCL Rituximab-CHOP Consortium Program Study.

Gene expression profiling (GEP) has stratified diffuse large B-cell lymphoma (DLBCL) into molecular subgroups that correspond to different stages of lymphocyte development-namely germinal center B-cell like and activated B-cell like. This classification has prognostic significance, but GEP is expensive and not readily applicable into daily practice, which has lead to immunohistochemical algorithms proposed as a surrogate for GEP analysis. We assembled tissue microarrays from 475 de novo DLBCL patients who were treated with rituximab-CHOP chemotherapy. All cases were successfully profiled by GEP on formalin-fixed, paraffin-embedded tissue samples. Sections were stained with antibodies reactive with CD10, GCET1, FOXP1, MUM1 and BCL6 and cases were classified following a rationale of sequential steps of differentiation of B cells. Cutoffs for each marker were obtained using receiver-operating characteristic curves, obviating the need for any arbitrary method. An algorithm based on the expression of CD10, FOXP1 and BCL6 was developed that had a simpler structure than other recently proposed algorithms and 92.6% concordance with GEP. In multivariate analysis, both the International Prognostic Index and our proposed algorithm were significant independent predictors of progression-free and overall survival. In conclusion, this algorithm effectively predicts prognosis of DLBCL patients matching GEP subgroups in the era of rituximab therapy.

IFN-gamma-independent autocrine cytokine regulatory mechanism in reprogramming of macrophage responses to bacterial lipopolysaccharide.

Macrophages are now well recognized to have a critical role in both innate and acquired immunity. The sentinel macrophage function is highly regulated and serves to allow for intrinsic plasticity of the innate immune responses to potential environmental signals. However, the mechanisms underlying the dynamic properties of the cellular arm of innate immunity are poorly understood. Therefore, we have conducted a series of in vitro studies to evaluate the contribution of immunoregulatory cytokines, such as IFN-gamma, IL-10, and IL-12, in modulation of macrophage responses. We found that macrophages from IFN-gamma knockout (IFN-gamma(-/-)) mice exhibit only marginal LPS-induced TNF-alpha, IL-12, and NO responses, all of which can be fully restored in the presence of rIFN-gamma. Pretreatment with substimulatory LPS concentrations led to reprogramming of IFN-gamma(-/-) macrophage responses in a dose-dependent manner that manifested by an increased TNF-alpha and IL-12, but not NO, production upon the subsequent LPS challenge. These reprogramming effects were substantially attenuated and profoundly enhanced in macrophages from IL-12(-/-) and IL-10(-/-) mice, respectively, as compared with those modulated in macrophages from the congenic wild-type mice. LPS-dependent reprogramming was also fully reproduced in macrophages isolated from SCID mice after immunodepletion of NK cells. Our data strongly imply that cytokine (TNF-alpha and IL-12), but not NO, responses in macrophages may, at least in part, be governed by an autocrine IFN-gamma-independent regulatory mechanism reciprocally controlled by IL-10 and IL-12. This mechanism may serve as an alternative/coherent pathway to the canonical IFN-gamma-dependent induction of antimicrobial and tumoricidal activity in macrophages.

Characteristics of salivary diffuse infiltrative lymphocytosis syndrome in West Africa.

OBJECTIVE: To determine the prevalence of diffuse infiltrative lymphocytosis syndrome (DILS) in the minor salivary glands of 30 African Cameroonian adults with the acquired immunodeficiency syndrome (AIDS). DESIGN: Salivary gland tissue was analyzed using a modified classification system that was developed to aid the diagnosis of Sjögren syndrome. The advantages and disadvantages of this approach are discussed. MATERIALS AND METHODS: Formalin-fixed, paraffin-embedded, hematoxylin-eosin-stained biopsy sections were prepared for 30 patients with AIDS, 26 healthy individuals who declined human immunodeficiency virus (HIV) testing, and 4 seronegative healthy controls. Tissues were immunostained for CD4/CD8+ lymphocytes and cytomegalovirus (CMV), and transmission electron microscopy was performed to locate viral particles. Patients were tested for HIV-1 and HIV-2 by the HIV/Chek System 3 or CAMSTIX-HIV-1 and HIV-2 assay. RESULTS: Severe salivary ductal atypia (96%) was the feature most strongly associated with AIDS, and the lymphocytic focus score was the second histologic feature most strongly correlated with AIDS. Forty-eight percent of patients with HIV-1 infection had more than 1 lymphocytic focus in a minor salivary gland. These lymphocytes were primarily CD8+. We report, to the best of our knowledge, the first case of multinucleated salivary duct epithelial cells in minor salivary glands also containing enveloped virus particles. All cases were negative for CMV. CONCLUSIONS: The prevalence of DILS in West Africans with AIDS appears higher than the prevalence reported in whites from the United States and Europe and in blacks from the United States, a group that has been reported to have a greater incidence of DILS than whites. This discrepancy may be related to differences in patient selection criteria. The determination of lymphocytic focus score, as used in the diagnosis of Sjögren syndrome, with the adjunct of ductal atypia is useful for assessing DILS. The impact of patient selection, drug therapy, and parasites on salivary gland pathology is discussed.

Immunohistochemical detection of cyclin D1 using optimized conditions is highly specific for mantle cell lymphoma and hairy cell leukemia.

Mantle cell lymphoma (MCL) is more aggressive when compared with other lymphomas composed of small, mature B lymphocytes. Cyclin D1 is overexpressed in MCL as a result of the translocation t(11;14)(q13;q32). Cyclin D1 immunohistochemistry in fixed, paraffin-embedded tissue contributes to the precise and reproducible diagnosis of MCL without the requirement of fresh tissue. However, its use in bone marrow biopsies is not well established. In addition, increased levels of cyclin D1 mRNA have been found in hairy cell leukemia but have not consistently been detected by immunohistochemistry. We used a polyclonal antibody and heat-induced antigen retrieval conditions to evaluate 73 fixed, paraffin-embedded bone marrow, spleen, and lymph node specimens with small B-cell infiltrates, obtained from 55 patients. Cyclin D1 was overexpressed in 13/13 specimens of MCL (usually strong, diffuse reactivity in most tumor cells) and in 14/14 specimens of hairy cell leukemia (usually weak, in a subpopulation of tumor cells). No reactivity was detected in five cases of B-chronic lymphocytic leukemia; five cases of splenic marginal zone lymphoma; six cases of nodal marginal zone cell lymphoma; two cases of gastric marginal zone cell lymphoma; or ten benign lymphoid infiltrates in bone marrow, spleen, or lymph nodes. In summary, although the total number of studied cases is small and a larger series of cases may be required to confirm our data, we present optimized immunohistochemical conditions for cyclin D1 in fixed, paraffin-embedded tissue that can be useful in distinguishing MCL and hairy cell leukemia from other small B-cell neoplasms and reactive lymphoid infiltrates.

Intravascular lymphomatosis with bone marrow involvement.

We describe the case of a 56-year-old man who presented with numbness and tingling of the extremities, weakness, and fatigue. Laboratory findings included anemia and thrombocytopenia. A diagnosis of intravascular lymphomatosis was established when liver, omentum, and bone marrow samples were examined. A review of the literature reveals that most cases of intravascular lymphomatosis have cytopenias, mainly anemia and thrombocytopenia, but bone marrow involvement is rare. In our case, a subtle neoplastic infiltrate in the marrow sinusoids was highlighted with a B-cell marker. While immunohistochemical analysis was not performed in most reported cases in the literature, our studies suggest that a systematic search in bone marrow of cases of intravascular lymphomatosis may reveal unsuspected neoplastic cells. We conclude that bone marrow involvement in intravascular lymphomatosis appears to be rare, has subtle features, and is difficult to diagnose if unsuspected and not searched for.

Immunocytochemical analysis of MNDA in tissue sections and sorted normal bone marrow cells documents expression only in maturing normal and neoplastic myelomonocytic cells and a subset of normal and neoplastic B lymphocytes.

The human myeloid cell nuclear differentiation antigen (MNDA) is a nuclear antigen known to be expressed in mature myelomonocytic cell lines. An extensive immunocytochemical evaluation of fixed tissues confirmed MNDA expression in normal maturing granulocytes and monocytes and in acute nonlymphocytic leukemias and chronic myelogenous leukemia. MNDA was not detected in normal tissue histiocytes but was found in activated macrophages and foreign body giant cells associated with inflammation. Flow cytometric cell sorting of normal bone marrow established that MNDA is initially expressed in myeloid blast cells. Examination of lymphoid tissues showed a low level of expression in a population of normal mande B lymphocytes but not in germinal center cells or plasma cells. A subset of B cell neoplasms expressing MNDA included hairy cell leukemia, parafollicular (monocytoid) B cell lymphoma, mantle cell lymphoma, and small lymphocytic lymphoma. Cell sorting of normal bone marrow showed MNDA expression in CD20+/CD10-/CD5- B cells. MNDA was not detected in other normal bone marrow or all other nonhematopoietic cells. The hematopoietic cell-specific pattern of MNDA expression was elucidated through a comprehensive analysis of normal and neoplastic tissues, and the results provide further evidence of the coexpression of B- and myeloid cell markers in neoplastic B cells and identify a normal B cell population that might be related to the cell of origin of a subset of B cell neoplasms.

Somatic mutation analysis of IgH variable regions reveals that tumor cells of most parafollicular (monocytoid) B-cell lymphoma, splenic marginal zone B-cell lymphoma, and some hairy cell leukemia are composed of memory B lymphocytes.

The cell of origin of parafollicular (monocytoid) B cell lymphoma (PBCL), splenic marginal zone lymphoma (SMZL), and hairy cell leukemia (HCL) is controversial. To better understand the relationship between these low-grade B-cell neoplasms, we analyzed the nucleotide sequences of the rearranged immunoglobulin heavy (IgH) chain variable (V) region of the clonal population of cells in five cases of PBCL, four cases of SMZL, and seven cases of HCL to determine whether these neoplasms could be differentiated by the degree of somatic mutation in the IgH V gene or by the IgH V gene family usage. DNA was extracted from diagnostic material and clonality confirmed by PCR. The DNA was reamplified using V heavy chain family specific primers, and the amplicons were sequenced. Sequences were compared with germline IgH V gene sequences, and base changes were determined to be silent or to represent amino acid replacements by using three different methods. Four of five (80%) cases of PBCL, three of four (75%) cases of SMZL, and three of seven (43%) cases of HCL showed evidence of antigen selection, suggesting that these neoplasms involved clonal expansions of postgerminal center memory lymphocytes. Only SMZL showed a preferential usage of V(H)1 family genes.

Chronic myeloid leukemia manifested during megakaryoblastic crisis.

We describe a 38-year-old man with a chronic myeloproliferative syndrome characterized by elevated white blood cell and platelet counts and increased blasts in the peripheral blood. Bone marrow aspiration was a "dry tap" and the biopsy specimen was hypercellular with numerous blasts, atypical megakaryocytes, and increased reticulin fibrosis. The blasts exhibited cytochemical reactivity for nonspecific esterase and PAS and immunohistochemically were positive for factor VIII, supporting megakaryoblastic lineage. Cytogenetic studies of peripheral blood revealed the t(9;22)(q34;q11). We interpreted these findings to be most consistent with chronic myeloid leukemia (CML) manifested at the time of megakaryoblastic crisis. Although the initial complete blood count showed leukocytosis and thrombocytosis, the patient subsequently had pancytopenia with clinical and pathologic findings consistent with acute myelofibrosis (AMF). Cytosine arabinoside and etoposide chemotherapy induced remission of the acute leukemia. We conclude that CML infrequently presents itself in megakaryoblastic crisis and that such cases may result in the clinicopathologic syndrome of AMF. The success of chemotherapy in this case also suggests that intensive antileukemic therapy may be useful in other patients with either CML-blast crisis or the clinicopathologic syndrome of AMF.

Stage IE non-Hodgkin's lymphoma involving the dura: A clinicopathologic study of five cases.

OBJECTIVE AND DESIGN: Non-Hodgkin's lymphomas rarely present as a localized mass involving the dura. In this report we describe the clinical, histologic, and immunohistochemical features of five cases of stage IE non-Hodgkin's lymphoma involving the dura. PATIENTS: Four women and one man, 36 to 67 years of age (median 50.6 years). RESULTS: Myelography and magnetic resonance imaging scans revealed discrete expansile masses involving the dura of the cervical, thoracic, and lumbar regions of the spinal cord and the frontal lobe of the brain. Histologically, the tumors were classified in the Working Formulation as small lymphocytic (2), diffuse large cell (2), and large cell immunoblastic (1) (anaplastic large cell lymphoma). Four tumors were of B-cell lineage and the anaplastic large-cell lymphoma was of T-cell lineage. The two small lymphocytic neoplasms had immunoglobulin heavy-chain gene rearrangements as shown by either Southern blot hybridization or the polymerase chain reaction. Four patients underwent decompression laminectomy; three received spinal radiation; two received chemotherapy (one intrathecal, one systemic) for lymphocytosis of the cerebrospinal fluid. The dural mass overlying the frontal lobe was excised and focally irradiated. Clinical follow-up was available for all patients. Four patients were alive 12 to 40 months after diagnosis and showed no evidence of recurrent or disseminated disease. The patient with anaplastic large-cell lymphoma died 10 days after laminectomy, secondary to pulmonary thromboemboli. CONCLUSIONS: We conclude that non-Hodgkin's lymphomas of varied histologic types and of either B- or T-cell lineage may rarely present as a stage IE dural mass. These lesions appear to have a good initial response to treatment; however, longer clinical follow-up is necessary to assess the incidence of relapse and final outcome.

Peritoneal silicosis.

A 73-year-old man with a clinical diagnosis of pulmonary silicosis (long-standing exposure to silica, pulmonary infiltrates, and flu-like symptoms) presented to the emergency room with fever, acute biliary colic, and cholelithiasis. The patient had a 2-year status postchemotherapy with complete remission of hepatic and splenic malignant lymphoma. At laparotomy we found studding of the undersurface of the diaphragm with multiple small dark nodules. Owing to the patient's history of previously treated abdominal malignant lymphoma, the lesions were grossly interpreted as abdominal lymphomatosis. The microscopic appearance of the lesions suggested silicotic nodules, which were confirmed by digital scanning electron microscopy and roentgenographic microanalysis performed on formalin-fixed, paraffin-embedded tissue. This is an unusual extrapulmonary pattern of peritoneal seeding in silicosis.