Whole-Body Imaging of Multiple Myeloma: Diagnostic Criteria.

Multiple myeloma (MM) is a clonal plasma cell proliferative disorder characterized by primary infiltration of bone marrow and excessive production of abnormal immunoglobulin. This disease is the second most common hematologic malignancy (after lymphoma), and its spectrum of characteristic features are widely known by the acronym CRAB (hypercalcemia, renal impairment, anemia, and bone lesions). Traditionally, the diagnosis and treatment of MM have been triggered by clear end-organ damage. However, owing to recently introduced treatment options that can extend patient survival and the increasing recognition of biomarkers that can be used to identify patients at high risk of progression to active disease, the diagnostic criteria have been revised. Bone disease is one of the most prominent features of MM, and imaging has an important role in diagnosis and follow-up, with each whole-body imaging modality having different indications in distinct disease situations. Skeletal survey has been the standard imaging procedure used during the past decade, but it should no longer be used unless it is the only option. Whole-body low-dose CT is a reasonable and cost-effective initial imaging approach. Whole-body MRI is the most sensitive technique for detecting bone involvement and assessing painful complications. PET/CT is the best tool for evaluating treatment response. The importance of radiologists has increased in this scenario. Therefore, to properly assist hematologists and improve the care of patients with MM, it is essential that radiologists know the updated diagnostic criteria for MM, indications for and limitations of each imaging option, and recommendations for follow-up. Online supplemental material is available for this article. ©RSNA, 2019.

CD200 has an important role in the differential diagnosis of mature B-cell neoplasms by multiparameter flow cytometry.

BACKGROUND: Multiparameter flow cytometry is a useful tool for the diagnostic evaluation of mature B-cell neoplasms (MBN). Recently, it has been shown that CD200 may improve the distinction between chronic lymphocytic leukemia (CLL; CD200+) and mantle cell lymphoma (MCL; CD200-), but the role of CD200 expression in atypical CLL and other MBN remains to be established. METHODS: To address this issue, we investigated the expression of CD200 in 159 consecutive cases of MBN. RESULTS: CD200 was strongly expressed in CLL and was revealed to be an excellent marker to distinguish CLL from MCL, even in cases of atypical CLL. However, lack of CD200 was not an exclusive finding of MCL, being also observed in other MBNs. Furthermore, CD200 was highly expressed in hairy cell leukemia, being useful in the differential diagnosis of lymphomas with villous lymphocytes. Herein, we propose an algorithm to classify CD5+ MBNs based on the expression of CD200, CD11c, heavy chain immunoglobulins, and Matutes score. CONCLUSIONS: These results expand the understanding of the CD200 expression in MBNs, giving further support for the inclusion of this marker in the routine investigation by flow cytometry.

CD200 has an important role in the differential diagnosis of mature B-cell neoplasms by multiparameter flow cytometry.

Background: Multiparameter flow cytometry is a useful tool for the diagnostic evaluation of mature B-cell neoplasms (MBN). Recently, it has been shown that CD200 may improve the distinction between chronic lymphocytic leukemia (CLL; CD200+) and mantle cell lymphoma (MCL; CD200-), but the role of CD200 expression in atypical CLL and other MBN remains to be established. Methods: To address this issue, we investigated the expression of CD200 in 159 consecutive cases of MBN. Results: CD200 was strongly expressed in CLL and was revealed to be an excellent marker to distinguish CLL from MCL, even in cases of atypical CLL. However, lack of CD200 was not an exclusive finding of MCL, being also observed in other MBNs. Furthermore, CD200 was highly expressed in hairy cell leukemia, being useful in the differential diagnosis of lymphomas with villous lymphocytes. Herein, we propose an algorithm to classify CD5+ MBNs based on the expression of CD200, CD11c, heavy chain immunoglobulins and Matutes score. Conclusions: These results expand the understanding of the CD200 expression in MBNs, giving further support for the inclusion of this marker in the routine investigation by flow cytometry. © 2013 Clinical Cytometry Society.

Serum free light chains and post-transplant lymphoproliferative disorder in patients with renal transplant.

The aim of the present study was to determine whether there is an association between serum free light chains (sFLC) quantification and the development of post-transplant lymphoproliferative disorder (PTLD), using serum samples from a nested case-control cohort of patients with renal transplant. Ten new cases of PTLD and 46 controls were enrolled. Additional comparison groups consisted of five human immunodeficiency virus (HIV)-infected individuals, five with untreated Hodgkin lymphoma and six normal individuals. Serum κ and λ FLC concentrations were measured by nephelometry and compared with reference ranges (normal and renal ranges). κ and/or λ were above the normal range in 90% of cases and in 65% of matched controls. There was no statistically significant difference between all groups, except for λ FLC concentrations between cases of PTLD and normal individuals (p = 0.016). The κ/λ sFLC ratios of cases and controls were within the renal range and normal range. Our results suggest that sFLC are not useful to predict PTLD development in renal transplant recipients.

Treatment-induced oxidative stress and cellular antioxidant capacity determine response to bortezomib in mantle cell lymphoma.

PURPOSE: Proteasome inhibition disrupts protein homeostasis and induces apoptosis. Up to 50% of patients with relapsed mantle cell lymphoma (MCL) respond to bortezomib. We used gene expression profiling to investigate the connection between proteasome inhibition, cellular response, and clinical efficacy. EXPERIMENTAL DESIGN: We assessed transcriptional changes in primary tumor cells from five patients during treatment with bortezomib in vivo, and in 10 MCL cell lines exposed to bortezomib in vitro, on Affymetrix microarrays. Key findings were confirmed by western blotting. RESULTS: MCL cell lines exposed to bortezomib in vitro showed upregulation of endoplasmic reticulum and oxidative stress response pathways. Gene expression changes were strongest in bortezomib-sensitive cells and these cells were also more sensitive to oxidative stress induced by H2O2. Purified tumor cells obtained at several timepoints during bortezomib treatment in 5 previously untreated patients with leukemic MCL showed strong activation of the antioxidant response controlled by NRF2. Unexpectedly, activation of this homeostatic program was significantly stronger in tumors with the best clinical response. Consistent with its proapoptotic function, we found upregulation of NOXA in circulating tumor cells of responding patients. In resistant cells, gene expression changes in response to bortezomib were limited and upregulation of NOXA was absent. Interestingly, at baseline, bortezomib-resistant cells displayed a relatively higher expression of the NRF2 gene-expression signature than sensitive cells (P < 0.001). CONCLUSION: Bortezomib triggers an oxidative stress response in vitro and in vivo. High cellular antioxidant capacity contributes to bortezomib resistance.

Bortezomib resistance in mantle cell lymphoma is associated with plasmacytic differentiation.

Bortezomib induces remissions in 30%-50% of patients with relapsed mantle cell lymphoma (MCL). Conversely, more than half of patients' tumors are intrinsically resistant to bortezomib. The molecular mechanism of resistance has not been defined. We generated a model of bortezomib-adapted subclones of the MCL cell lines JEKO and HBL2 that were 40- to 80-fold less sensitive to bortezomib than the parental cells. Acquisition of bortezomib resistance was gradual and reversible. Bortezomib-adapted subclones showed increased proteasome activity and tolerated lower proteasome capacity than the parental lines. Using gene expression profiling, we discovered that bortezomib resistance was associated with plasmacytic differentiation, including up-regulation of IRF4 and CD38 and expression of CD138. In contrast to plasma cells, plasmacytic MCL cells did not increase immunoglobulin secretion. Intrinsically bortezomib-resistant MCL cell lines and primary tumor cells from MCL patients with inferior clinical response to bortezomib also expressed plasmacytic features. Knockdown of IRF4 was toxic for the subset of MCL cells with plasmacytic differentiation, but only slightly sensitized cells to bortezomib. We conclude that plasmacytic differentiation in the absence of an increased secretory load can enable cells to withstand the stress of proteasome inhibition. Expression of CD38 and IRF4 could serve as markers of bortezomib resistance in MCL. This study has been registered at http://clinicaltrials.gov as NCT00131976.

SAGE analysis demonstrates increased expression of TOSO contributing to Fas-mediated resistance in CLL.

To identify novel genes involved in the molecular pathogenesis of chronic lymphocytic leukemia (CLL) we performed a serial analysis of gene expression (SAGE) in CLL cells, and compared this with healthy B cells (nCD19(+)). We found a high level of similarity among CLL subtypes, but a comparison of CLL versus nCD19(+) libraries revealed 55 genes that were over-represented and 49 genes that were down-regulated in CLL. A gene ontology analysis revealed that TOSO, which plays a functional role upstream of Fas extrinsic apoptosis pathway, was over-expressed in CLL cells. This finding was confirmed by real-time reverse transcription-polymerase chain reaction in 78 CLL and 12 nCD19(+) cases (P < .001). We validated expression using flow cytometry and tissue microarray and demonstrated a 5.6-fold increase of TOSO protein in circulating CLL cells (P = .013) and lymph nodes (P = .006). Our SAGE results have demonstrated that TOSO is a novel over-expressed antiapoptotic gene in CLL.

Flow cytometry characterization of leukemic phase of nasal NK/T-cell lymphoma in tumor biopsies and peripheral blood.

We report the findings of the immunophenotypic profile of three cases of nasal T/NK cell lymphoma in leukemic phase. Flow cytometry analysis was carried out using cell suspensions of tumor nasal biopsies and peripheral blood. Tumor samples were composed by a mixture of a predominant subset of medium-size true NK cytCD3epsilon-, sCD3epsilon-, CD56+ cells mixed with a minor subset of medium-size T/NK sCD3epsilon+, CD56+ cells. Both subsets were also detected in peripheral blood. In addition, an infiltration of small-size sCD3epsilon+, CD56- normal T lymphocytes was also present.

PRAME is a membrane and cytoplasmic protein aberrantly expressed in chronic lymphocytic leukemia and mantle cell lymphoma.

The preferentially expressed antigen in melanoma (PRAME) gene is aberrantly expressed in chronic lymphoproliferative disorders (CLD). We produced and characterized an anti-PRAME monoclonal antibody (MoAb), which was then applied in a quantitative flow cytometric (QFC) method to evaluate PRAME expression in leukemic cells from the peripheral blood (PB) of 47 patients with chronic lymphocytic leukemia and seven with mantle cell lymphoma as well as in the PB mononuclear cells (PBMCs) and B lymphocytes from 15 healthy subjects. Approximately 90% of CLD, but none of the normal samples, presented more than 20% of PRAME+ lymphocytes. Moreover, the intensity of PRAME expression was significantly higher in CLD cells compared to normal B lymphocytes and PBMCs. By immunofluorescence microscopy and by permeabilized flow cytometry we demonstrated that PRAME is a membrane antigen and a cytoplasmic protein aberrantly expressed in malignant CLD. Our results suggest that the analysis of PRAME protein may contribute for the distinction between normal and leukemic cells in CLD, and that PRAME may be a potential target for therapy.

Mesenchymal stem cells can be obtained from the human saphena vein.

Mesenchymal stem cells (MSC) can be isolated from many sites adults and the fetus. Cells with osteoblastic, chondrogenic, leiomiogenic and stromogenic potentials have been obtained from the bovine artery wall, and we now show that MSC can be isolated also from the adult human vein wall. Cells detached from internal surface of the saphenous vein are cultured in vitro for 2-3 weeks and replated weekly. The culture forms a semi-confluent layer of spindle-shaped cells that are CD13(+), CD29(+), CD44(+), CD34(-), CD45(-), CD14(-), CD133(-), CD31(-), CD33(-), CD54(+), CD106(-), CD90(+), KDR(-), cadherin-5-, HLA class I(+) and HLA-DR- and differentiate in vitro into osteoblasts, chondrocytes and adipocytes. Gene expression, when compared with seven other normal tissues, shows strong similarity with MSC obtained from other sources. Three genes more expressed in saphenous MSC than in the other two MSC are related to angiogenesis, and the expression of two of them is shared by endothelial cells. These results demonstrate that the human vein wall contains mesenchymal cells with morphologic features, immunophenotypic markers, gene expression profile and differentiation potential that are similar to MSC obtained from the bone marrow and from the umbilical vein.

[Hematological features and expression profile of myeloid antigens of acute promyelocytic leukemia patients: analysis of prognostic factors for development of the retinoic acid syndrome]. / Características hematológicas e perfil de expressão de antígenos mielóides de pacientes com leucemia promielocítica aguda: análise de fatores prognósticos para o desenvolvimento da síndrome do ácido retinóico.

BACKGROUND: Acute Promyelocytic Leukemia (APL) is characterized by its good response to treatment with all trans retinoic acid (ATRA). However, some patients receiving ATRA develop a serious complication called retinoid syndrome (RS). The objective of this study was to compare the hematological and immunophenotypic features of APL patients who developed RS with those who did not. METHODS: We reviewed the medical records, roentgenograms, peripheral blood smears and bone marrow aspirates from 71 APL patients. Immunophenotypic analyses were available in 56 of these cases. Eight cases of RS were detected, whose clinical presentation was characterized by respiratory distress (n = 8), impairment of the renal function (n = 2), fever (n = 5), weight gain (n = 3), edema (n = 3) and/or pleural effusion (n = 5). The following variables were compared in patients with and without RS: hemoglobin levels, leukocyte and platelet counts, frequency of hypergranular and variant morphological subtypes, percentages of CD33+, CD13+, CD117+ blasts in the bone marrow, fluorescence intensity and variation of these markers in the leukemic cells, expressed as the median channel of fluorescence (MCF) and fluorescence coefficient of variation (CV), respectively. RESULTS: RS incidence was 11.26% and the average time for syndrome development was 11.5 days after starting ATRA treatment. All patients presented acute respiratory distress. Other symptoms included fever, weight gain, edema and renal insufficiency. The main radiological findings were ground glass opacities, increased vascular pedicle and peribronchial cuffing. There was no significant correlation between the variables selected and the risk of development of RS, however the Odds Ratios for patients presenting MCF for CD117 > 30 ua and CV for CD33 < 50 were of 7.14 (P = 0.08) and 7.86 (P = 0.06), respectively. CONCLUSIONS: The incidence, as well as the clinical, radiological and laboratory features of RS in this group of Brazilian APL patients were similar to those described in literature. None of the variables studied were significantly correlated to a higher risk of developing RS.

Características hematológicas e perfil de expressão de antígenos mielóides de pacientes com leucemia promielocítica aguda: análise de fatores prognósticos para o desenvolvimento da síndrome do ácido retinóico / Hematological features and expression profile of myeloid antigens of acute promyelocytic leukemia patients: analysis of prognostic factors for development of the retinoic acid syndrome

OBJETIVO: A leucemia promielocítica aguda (LPA) apresenta uma boa resposta ao tratamento com o ácido all trans retinóico (ATRA). Entretanto, alguns pacientes desenvolvem uma complicação grave chamada síndrome do ácido retinóico (SAR). O objetivo deste estudo foi comparar as características hematológicas e imunofenotípicas de pacientes com LPA que desenvolveram a SAR com as daqueles que não a desenvolveram. MÉTODOS: Foram analisados retrospectivamente os prontuários, exames radiológicos, lâminas de esfregaço de sangue e medula óssea de 71 pacientes com LPA, dos quais a análise imunofenotípica havia sido realizada em 56 casos. Foram identificados oito casos de SAR que, do ponto de vista clínico, caracterizaram-se por insuficiência respiratória (n=8), insuficiência renal (n=2), febre (n=5), ganho ponderal (n=3), edema periférico (n=3) e derrame pleural (n=5). As seguintes variáveis foram comparadas entre pacientes com e sem SAR: dosagem de hemoglobina, contagens de leucócitos e plaquetas no sangue periférico, distribuição dos subtipos hipergranular e variante, percentagens de blastos CD33+, CD13+, CD117+ na medula óssea, intensidade e variação dos valores de fluorescência destes antígenos nas células leucêmicas, expressas através dos canais medianos (CMFs) e dos coeficientes de variação (CVs) de fluorescência, respectivamente. RESULTADOS: A incidência da SAR foi de 11,26 por cento e o tempo médio para seu desenvolvimento 11,5 dias do início do tratamento. Todos os pacientes apresentaram desconforto respiratório agudo, por vezes associado à febre, ganho de peso, edema e insuficiência renal. Os achados radiológicos mais comuns foram: opacidades em vidro fosco, derrame pleural, espessamento peribrônquico e aumento da trama vascular pulmonar. Nenhuma das variáveis laboratoriais analisadas correlacionou-se significativamente ao risco de desenvolvimento da SAR, entretanto as Odd Ratios para CMF para o CD117 > 30 ua e CV para o CD33 < 50 foram de 7,14 (P=0,08) e de 7,86 (P=0,06), respectivamente. CONCLUSAO: A incidência e as características da SAR neste grupo de pacientes brasileiros foi semelhante à descrita na literatura. Nenhum dos parâmetros estudados correlacionou-se significativamente a um maior risco de desenvolvimento desta complicação.

Microgranular and t(11;17)/PLZF-RARalpha variants of acute promyelocytic leukemia also present the flow cytometric pattern of CD13, CD34, and CD15 expression characteristic of PML-RARalpha gene rearrangement.

Acute promyelocytic leukemia (APL) is a subtype acute myeloid leukemia in which leukemic promyelocytes predominate in the bone marrow (BM). Rapid diagnosis is critical for treatment decision since all-trans-retinoic acid must be administrated promptly. The microgranular variant may be of difficult diagnosis, as it may be confused with other diseases on morphological grounds. The purpose of this study was to determine if the microgranular variant has the same antigenic profile as the classical hypergranular type. The immunophenotype of leukemic cells from the bone marrow of 50 patients, with the PML-RARalpha gene rearrangement confirmed by RT-PCR, was determined by flow cytometry using a large panel of 22 monoclonal antibodies and a polyclonal anti-TdT antibody. Thirty-four cases were classified as classical APL and 16 as microgranular APL. The immunophenotypic profile of the two subtypes was indistinguishable concerning the presence or absence of these antigens, including the absence of reactivity for the HLA-DR antigen. The simultaneous immunophenotypic combination of a unique major cell population, heterogeneous intensity of expression of CD13, and the typical pattern of CD15/CD34 expression were similarly present in the hypergranular and microgranular subtypes. Homogeneous expression of CD33 was observed in 76% of the classical APL cases and in 100% of the microgranular cases. Additionally, we have studied two cases of PLZF-RARalpha APL that also displayed the same immunophenotype described for classical APL. Thus, the immunophenotypic profile highly characteristic of the PML-RARalpha gene rearrangement was also observed in microgranular and PLZF-RARalpha variants of APL.

Expression of CD117 and CD11b in bone marrow can differentiate acute promyelocytic leukemia from recovering benign myeloid proliferation.

The morphologic characteristics of bone marrow aspirates from patients recovering from acute agranulocytosis may be closely similar to the pattern observed in cases of acute promyelocytic leukemia (APL). The clinical manifestation also can be ambiguous in a substantial number of cases. The immunophenotypic features of bone marrow from 5 patients recovering from acute agranulocytosis, showing an increase in the percentage of promyelocytes (26%-66%), were compared with the immunophenotype of 31 consecutive patients with APL whose diagnosis was confirmed by PML-RAR alpha gene rearrangement. All markers were similarly expressed, except for CD117 and CD11b. CD117 was positive in 24 (77%) of the APL cases and in none of the acute agranulocytosis cases. On the other hand, CD11b was positive in 5 (100%) of the acute agranulocytosis cases and in only 2 (6%) of the APL cases. Thus, the CD117-CD11b+ phenotype was detected in all patients recovering from agranulocytosis and in only 1 (3%) of 31 APL cases. Therefore, we suggest that the combination of both markers is helpful in the differentiation of APL from recovering benign myeloid proliferation.