Cristiane Bedran Milito

CD10 and Bcl-2 expression combined with the International Prognostic Index can identify subgroups of patients with diffuse large-cell lymphoma with very good or very poor prognoses

Aims : Diffuse large B‐cell lymphoma (DLBCL) is characterized by marked biological heterogeneity. The identification of reproducible parameters that can be combined with the International Prognostic Index (IPI) to better predict outcome could lead to the development of effective risk‐adaptive strategies.

Contribution of Multiparameter Flow Cytometry Immunophenotyping to the Diagnostic Screening and Classification of Pediatric Cancer

Pediatric cancer is a relatively rare and heterogeneous group of hematological and non-hematological malignancies which require multiple procedures for its diagnostic screening and classification. Until now, flow cytometry (FC) has not been systematically applied to the diagnostic work-up of such malignancies, particularly for solid tumors. Here we evaluated a FC panel of markers for the diagnostic screening of pediatric cancer and further classification of pediatric solid tumors. The proposed strategy aims at the differential diagnosis between tumoral vs. reactive samples, and hematological vs. non-hematological malignancies, and the subclassification of solid tumors. In total, 52 samples from 40 patients suspicious of containing tumor cells were analyzed by FC in parallel to conventional diagnostic procedures. The overall concordance rate between both approaches was of 96% (50/52 diagnostic samples), with 100% agreement for all reactive/inflammatory and non-infiltrated samples as well as for those corresponding to solid tumors (n = 35), with only two false negative cases diagnosed with Hodgkin lymphoma and anaplastic lymphoma, respectively. Moreover, clear discrimination between samples infiltrated by hematopoietic vs. non-hematopoietic tumor cells was systematically achieved. Distinct subtypes of solid tumors showed different protein expression profiles, allowing for the differential diagnosis of neuroblastoma (CD56hi/GD2+/CD81hi), primitive neuroectodermal tumors (CD271hi/CD99+), Wilms tumors (>1 cell population), rhabdomyosarcoma (nuMYOD1+/numyogenin+), carcinomas (CD45−/EpCAM+), germ cell tumors (CD56+/CD45−/NG2+/CD10+) and eventually also hemangiopericytomas (CD45−/CD34+). In summary, our results show that multiparameter FC provides fast and useful complementary data to routine histopathology for the diagnostic screening and classification of pediatric cancer.

The prognostic value of the expression of Bcl-2, p53 and LMP-1 in patients with Hodgkin's lymphoma.

This study was undertaken to evaluate the clinical significance of the expression of Bcl-2, p53 and LMP-1 in Hodgkin and Reed - Sternberg cells of patients with Hodgkins lymphoma. The expression of these proteins in pre-treatment tissue biopsy specimens was correlated with presenting clinical features, failure-free survival (FFS) and overall survival (OS) in 83 patients with a confirmed Hodgkins lymphoma treated in a single institution. HIV-positive patients were excluded. Patients were classified according to the International Prognostic Score (IPS) in low-risk (0 - 2 factors) and high-risk groups. The median age was 41 years (15 - 84), 41% were women, and 93% had advanced-stage disease (IIB - IVB). The expression of Bcl-2, p53 and LMP-1 was not associated with the complete remission rate, FFS or OS. The IPS risk group was the only factor significantly associated with OS. Patients with a high IPS had a lower 5 year OS (43% vs. 79%, P = 0.003). The expression of Bcl-2, p53 and LMP-1 did not add prognostic information to the IPS.

PKC-beta II expression has prognostic impact in nodal diffuse large B-cell lymphoma.

Recent studies of gene expression and immunohistochemistry have shown that protein kinase C-beta II (PKC-beta II) might have prognostic significance in patients with diffuse large B-cell lymphoma (DLBCL). We sought to determine the prognostic significance of the expression of PKC-beta II in patients with nodal DLBCL. Formalin-fixed, paraffin-embedded tissues were stained with a monoclonal antibody to PKC-beta II protein. A total of 125 patients were studied; 83 patients (66%) were in the low-risk International Prognostic Index (IPI) group. Forty-eight patients (38%) were positive for PKC-beta II. Complete remission was obtained in 70%, and was not influenced by the PKC-beta II status (67 vs 71%). The 5-year event-free survival (EFS) was worse in high-risk patients (14 vs 58%, P<0.001) and in those with PKC-beta II positivity (36 vs 49%, P=0.054). In low-risk IPI patients, PKC-beta II expression was related to a worse 5-year overall survival (OS) (60 vs 76%, P=0.033) and a worse 5-year EFS (48 vs 66%, P=0.014). In a Cox regression analysis for EFS, both PKC-beta II expression (hazard ratio=1.68, P=0.037) and the IPI (HR=3.07, P<0.001) were independent poor prognostic factors. PKC-beta II (HR=1.72, P=0.046) and the IPI (HR=5.16, P<0.001) were also independent poor prognostic factors for the OS. PKC-beta II expression, along with the IPI, were associated with a worse EFS and OS in patients with nodal DLBCL specially in low-risk IPI patients.

Nodal and Extranodal Plasmacytomas Expressing Immunoglobulin A: An Indolent Lymphoproliferative Disorder With a Low Risk of Clinical Progression

Plasmacytomas expressing immunoglobulin A are rare and not well characterized. In this study, 9 cases of IgA-positive plasmacytoma presenting in lymph node and 3 in extranodal sites were analyzed by morphology, immunohistochemistry, and polymerase chain reaction examination of immunoglobulin heavy and κ light chain genes. Laboratory features were correlated with clinical findings. There were 7 males and 5 females; age range was 10 to 66 years (median, 32 y). Six of the patients were younger than 30 years of age, 5 of whom had nodal disease. About 67% (6 of 9) of the patients with nodal disease had evidence of immune system dysfunction, including human immunodeficiency virus infection, T-cell deficiency, autoantibodies, arthritis, Sjögren syndrome, and decreased B cells. An IgA M-spike was detected in 6 of 11 cases, and the M-protein was nearly always less than 30 g/L. All patients had an indolent clinical course without progression to plasma cell myeloma. Histologically, nodal IgA plasmacytomas showed an interfollicular or diffuse pattern of plasma cell infiltration. The plasma cells were generally of mature Marschalko type with little or mild pleomorphism and exclusive expression of monotypic IgA. There was an equal expression of κ and λ light chains (ratio 6:6). Clonality was showed in 9 of 12 cases: by polymerase chain reaction in 7 cases, by cytogenetic analysis in 1 case, and by immunofixation in 1 case. Clonality did not correlate with pattern of lymph node infiltration. Our results suggest that IgA plasmacytomas may represent a distinct form of extramedullary plasmacytoma characterized by younger age at presentation, frequent lymph node involvement, and low risk of progression to plasma cell myeloma.

Evaluation of intra- and interobserver agreement and its clinical significance for scoring bcl-2 immunohistochemical expression in diffuse large B-cell lymphoma

Immunohistochemistry (IHC) has become an essential part of diagnosis and clinical research in lymphomas. There is considerable heterogeneity, however, in IHC findings regarding expression rate and positivity cut‐offs, which creates a degree of uncertainty that has prevented its incorporation for prognostic purposes. The purpose of the present study was to assess intra‐ and interobserver agreement in scoring bcl‐2 expression on IHC. The study materials were 81 diffuse large B‐cell lymphomas. Slides were processed in the same laboratory, and were analyzed independently and in a blinded manner by four pathologists twice, at least 1 month apart. The positivity rates ranged from 31% to 41% in the first evaluation, and from 30% to 43% in the second evaluation. The two analyses by the same pathologist gave concordant results in 88–93% of cases (κ = 0.71–0.83). Complete agreement among all observers varied from 72% to 79%. The experience of the observer did not influence intra‐observer concordance. Cooperative analysis of discordant slides led to consensus in all cases. The variation observed in scoring bcl‐2 expression is acceptable for use in lymphoma diagnosis and classification. The use of IHC stratification, however, for clinical decisions regarding treatment will require standardization and centralized consensus review, and must await the results of ongoing prospective trials.

Classificação dos linfomas não-Hodgkin: estudo morfológico e imunoistoquímico de 145 casos

The non-Hodgkins lymphomas classifications have been a controversial reason for the last thirty years. Many classifications have been proposed trying to achieve a consensus among pathologists and clinicians. The objective of this study was to analyse critically three of these classifications by the retrospective study of 145 cases of lymph nodes primary lymphomas, selected from the department of Pathology of the Hospital Universitario Clementino Fraga Filho between 1979 and 1995. The revised cases were classifiedby the proposal of Working Formulation, Kiel and Real. Immunohistochemical detections were employed with antibodies anti-CD45, anti-CD20, anti-CD45RO and anti-CD30. One hundred and seven cases (73.7%) showed B phenotype; thirty three cases (22.7%), T phenotype and 4 cases were null (anaplastic large cell lymphomas). It was possible to predict the phenotype by the morphology in 89,4% of the cases.The high grade lymphomas predominated (59.2%), and the centroblastic lymphoma was more frequent (31.7%). The follicular lymphomas account for 29 cases (20%), the large cell type (31%) have more incidence than the small cell type (20%). Comparing the three classifications, we observed that certain Working Formulations groups agglomerate multiple entities. This occurs because it is based only on morphologic characteristics, hence should not be used. Kiel and Real classifications should have their employ stimulated because they present a good histopathologic analysis, a immunologic study that defines biologic entities correlating, whenever possible, with the postulated normal counterpart.

Treatment outcomes for Hodgkin lymphoma: First report from the Brazilian Prospective Registry

Data about Hodgkin lymphoma (HL) in developing countries are scarce and suggest the existence of substantial disparities in healthcare and outcomes in large areas of the world. In 2009, a prospective registry of HL was implemented in Brazil. Web‐based data were contributed by 20 institutions across the country participating in the Brazilian Prospective Hodgkins Lymphoma Registry. The aim of this study was to present the clinical features and outcomes of newly diagnosed patients with HL aged 13 to 90 years. Multivariate Cox regression models were used to estimate progression‐free (PFS) and overall survival (OS) by clinical factors. A total of 674 patients with classical HL were analysed, with a median follow‐up of 37 months. Median age was 30 years (13‐90). The median time from the onset of symptoms to diagnosis was 6 months (0‐60). Only 6% of patients had early favourable disease, while 65% had advanced disease. Stage IVB was present in 26% and a high‐risk International Prognostic Score in 38%. Doxorubicin, bleomycin, vinblastine, and dacarbazine was used in 93%. The median dose of radiotherapy was 36 Gy for localized disease and 32 Gy for advanced disease. The 3 year PFS in early favourable, early unfavourable, and advanced disease were 95%, 88%, and 66%, respectively. High‐risk International Prognostic Score, advanced disease, and age greater than or equal to 60 were independently associated with poorer PFS and OS; performance status greater than or equal to 2 was also associated with a poorer OS. Poor‐risk patients predominated. Radiation doses for localized disease appear higher than current recommendations. Outcomes appear inferior in developing countries than in developed countries. Delayed diagnosis is probably a major factor underlying these findings. Scattered reports from developing nations suggest that many aspects of standard care in developed countries remain unmet needs for populations living in developing countries. The present report contributes to this body of data, with a proper description of what is currently achieved in urban areas in Brazil.

DUOX1 Silencing in Mammary Cell Alters the Response to Genotoxic Stress

DUOX1 is an H2O2-generating enzyme related to a wide range of biological features, such as hormone synthesis, host defense, cellular proliferation, and fertilization. DUOX1 is frequently downregulated in lung and liver cancers, suggesting a tumor suppressor role for this enzyme. Here, we show that DUOX1 expression is decreased in breast cancer cell lines and also in breast cancers when compared to the nontumor counterpart. In order to address the role of DUOX1 in breast cells, we stably knocked down the expression of DUOX1 in nontumor mammary cells (MCF12A) with shRNA. This led to higher cell proliferation rates and decreased migration and adhesion properties, which are typical features for transformed cells. After genotoxic stress induced by doxorubicin, DUOX1-silenced cells showed reduced IL-6 and IL-8 secretion and increased apoptosis levels. Furthermore, the cell proliferation rate was higher in DUOX1-silenced cells after doxorubicin medication in comparison to control cells. In conclusion, we demonstrate here that DUOX1 is silenced in breast cancer, which seems to be involved in breast carcinogenesis.

Lower socioeconomic status is independently associated with shorter survival in Hodgkin Lymphoma patients—An analysis from the Brazilian Hodgkin Lymphoma Registry

Socioeconomic status (SES) is a well‐known determinant of outcomes in cancer. The purpose of this study was to analyze the impact of the SES on the outcomes of Hodgkin lymphoma (HL) patients from the Brazilian Prospective HL Registry. SES stratification was done using an individual asset/education‐based household index. A total of 624 classical HL patients with diagnosis from January/2009 to December/2014, and treated with ABVD (doxorubicin, bleomycin, vinblastine and dacarbazine), were analyzed. The median follow‐up was 35.6 months, and 33% were classified as lower SES. The 3‐year progression‐ free survival (PFS) in higher and lower SES were 78 and 64% (p < 0.0001), respectively. The 3‐year overall survival (OS) in higher and lower SES were 94 and 82% (p < 0.0001), respectively. Lower SES patients were more likely to be ≥ 60 years (16 vs. 8%, p = 0.003), and to present higher risk International Prognostic score (IPS) (44 vs. 31%, p = 0.004) and advanced disease (71 vs. 58%, p = 0.003). After adjustments for potential confounders, lower SES remained independently associated with poorer survival (HR = 3.12 [1.86‐5.22] for OS and HR = 1.66 [1.19‐2.32] for PFS). The fatality ratio during treatment was 7.5 and 1.3% for lower and higher SES (p = 0.0001). Infections and treatment toxicity accounted for 81% of these deaths. SES is an independent factor associated with shorter survival in HL in Brazil. Potential underlying mechanisms associated with the impact of SES are delayed diagnosis and poorer education. Educational and socio‐economic support interventions must be tested in this vulnerable population.