Dalton Alencar Fisher Chamone

p63 Protein expression in high risk diffuse large B-cell lymphoma

Background: p63 gene is a p53 homologue that encodes proteins with transactivation, DNA-binding and tetramerisation domains. The isoforms TAp63 and TAp73 transactivate p53 target genes and induce apoptosis, whereas the isoforms ΔNp63 and ΔNp73 lack transactivation and might have dominant-negative effects in p53 family members. p63 is expressed in germinal centre lymphocytes and can be related to the development of the lymphoma, but the prognostic significance of its expression in the survival of patients with diffuse large B-cell lymphoma (DLBCL) remains unclear. Aims: To determine whether quantitative immunohistochemical (IHC) analysis of p63 protein expression correlates with CD10 antigen, Bcl-6 antigen and IRF4 antigen expression and to determine whether p63 is a surrogate predictor of overall survival in high–intermediate and high risk DLBCL populations. Methods: CD10, Bcl-6 and IRF4 expression were retrospectively evaluated by IHC in 73 samples of high–intermediate and high risk DLBCL and were used to divide the lymphomas into subgroups of germinal centre B-cell-like (GCB) and activate B-cell-like (ABC) DLBCL. Similarly, p63 expression was evaluated by IHC and the results were compared with subgroups of DLBCL origin and with the survival rates for these patients. Results: p63 was expressed in more than 50% of malignant cells in 11 patients and did not show correlation with subgroups of GCB-like DLBCL or ABC-like DLBCL, but p63(+) patients had better disease-free survival (DFS) than those who were negative (pu200a=u200a0.01). Conclusions: p63(+) high–intermediate and high risk DLBCL patients have a better DFS than negative cases.

Effect of combined administration of saruplase and single-chain alteplase on coronary recanalization in acute myocardial infarction

Abstract The effectiveness of recombinant single- or 2-chain tissue-type plasminogen activator (rt-PA, alteplase) l and single-chain glycosylated* and unglycosylated 3–5 urokinase-type plasminogen activator in promoting the recanalization of thrombosed coronary arteries in patients with acute myocardial infarction is well established. Because the intrinsic fibrin selectivity of these 2 plasminogen activators is mediated by different molecular mechanisms, 6,7 their combined effect on clot dissolution might be more than additive. Combinations of these drugs in animal models of thrombosis produced significantly greater lysis than could be explained on the basis of their additive effects. 8 In a pilot study in myocardial infarction, a relatively low dose of 2-chain alteplase and single-chain urokinase-type plasminogen activator given together induced recanalization of the occluded coronary in almost all patients. 9,10 We designed this trial to attempt to confirm these preliminary findings. Low doses of single-chain rt-PA and unglycosylated single-chain urokinase-type plasminogen activator (saruplase)-corresponding to approximately 20% and between 12 to 25%, respectively, of the currently recommended dose of each thrombolytic drug—were used.

Quantification of imatinib in human serum: validation of a high-performance liquid chromatography-mass spectrometry method for therapeutic drug monitoring and pharmacokinetic assays

Background Imatinib mesylate has been a breakthrough treatment for chronic myeloid leukemia. It has become the ideal tyrosine kinase inhibitor and the standard treatment for chronic-phase leukemia. Striking results have recently been reported, but intolerance to imatinib and noncompliance with treatment remain to be solved. Molecular monitoring by quantitative real-time polymerase chain reaction is the gold standard for monitoring patients, and imatinib blood levels have also become an important tool for monitoring. Methods A fast and cheap method was developed and validated using high-performance liquid chromatography-mass spectrometry for quantification of imatinib in human serum and tamsulosin as the internal standard. Remarkable advantages of the method includes use of serum instead of plasma, less time spent on processing and analysis, simpler procedures, and requiring reduced amounts of biological material, solvents, and reagents. Stability of the analyte was also studied. This research also intended to drive the validation scheme in clinical centers. The method was validated according to the requirements of the US Food and Drug Administration and Brazilian National Health Surveillance Agency within the range of 0.500–10.0 μg/mL with a limit of detection of 0.155 μg/mL. Stability data for the analyte are also presented. Conclusion Given that the validated method has proved to be linear, accurate, precise, and robust, it is suitable for pharmacokinetic assays, such as bioavailability and bioequivalence, and is being successfully applied in routine therapeutic drug monitoring in the hospital service.

Determination of serum levels of imatinib mesylate in patients with chronic myeloid leukemia: validation and application of a new analytical method to monitor treatment compliance

Objective The goal of this study was to monitor imatinib mesylate therapeutically in the Tumor Biology Laboratory, Department of Hematology and Hemotherapy, Hospital das Clínicas, Faculdade de Medicina, Universidade de São Paulo (USP). A simple and sensitive method to quantify imatinib and its metabolite (CGP74588) in human serum was developed and fully validated in order to monitor treatment compliance. Methods The method used to quantify these compounds in serum included protein precipitation extraction followed by instrumental analysis using high performance liquid chromatography coupled with mass spectrometry. The method was validated for several parameters, including selectivity, precision, accuracy, recovery and linearity. Results The parameters evaluated during the validation stage exhibited satisfactory results based on the Food and Drug Administration and the Brazilian Health Surveillance Agency (ANVISA) guidelines for validating bioanalytical methods. These parameters also showed a linear correlation greater than 0.99 for the concentration range between 0.500 µg/mL and 10.0 µg/mL and a total analysis time of 13 minutes per sample. This study includes results (imatinib serum concentrations) for 308 samples from patients being treated with imatinib mesylate. Conclusion The method developed in this study was successfully validated and is being efficiently used to measure imatinib concentrations in samples from chronic myeloid leukemia patients to check treatment compliance. The imatinib serum levels of patients achieving a major molecular response were significantly higher than those of patients who did not achieve this result. These results are thus consistent with published reports concerning other populations.

Results of CHOP chemotherapy for diffuse large B-cell lymphoma

Patients with diffuse large B-cell lymphoma treated in a University Hospital were studied from 1990 to 2001. Two treatment regimens were used: ProMACE-CytaBOM and then, from November 1996 on, the CHOP regimen. Complete remission (CR), disease-free survival (DFS), and overall survival (OS) rates were determined. Primary refractory patients and relapsed patients were also assessed. A total of 111 patients under 60 years of age were assessed and ranked according to the international prognostic index adjusted to age. Twenty (18%) of them were classified as low risk, 40 (36%) as intermediate risk, 33 (29.7%) as high intermediate risk, and 18 (16.3%) as high risk. Over a five-year period, OS and DFS rates were 71 and 59%, respectively, for all patients. For the same time period, OS and DFS rates were 72.8 and 61.3%, respectively, for 77 patients treated with CHOP chemotherapy and 71.3 and 60% for patients treated with the ProMACE-CytaBOM protocol. There was no significant difference in OS or DFS between the two groups. Eleven of 50 refractory and relapsed patients were consolidated with high doses of chemotherapy. Three received allogenic and 8 autologous bone marrow transplantation. For the latter, CR was 62.5% and mean OS was 41.1 months. The clinical behavior, CR, DFS, and OS of the present patients were similar to those reported in the literature. We conclude that both the CHOP and ProMACE-CytaBOM protocols can be used to treat diffuse large B-cell lymphoma patients, although the CHOP protocol is preferable because of its lower cost and lower toxicity.

Prognostic impact of diffuse large B-cell lymphoma subgroups in patients undergoing autologous SCT

A total of 53 patients aged 18–60 years with high-intermediate or high-risk diffuse large B-cell lymphoma (DLBCL) were evaluated to analyze the impact of the cell of origin. Of 53 patients, 16 underwent autologous SCT (ASCT) in first remission and the rest received conventional chemotherapy. Immunohistochemistry was evaluated in 47 cases 17 were of germinal center (GC) origin and 30 were of non-GC origin. There was no survival difference between the two groups. Overall survival (OS) and disease-free survival (DFS) at 3 years were 93 and 83%, respectively, for the 14 patients who underwent ASCT. Their DFS was significantly better than that of patients who achieved CR but did not undergo ASCT. We conclude that ASCT is safe and improves the DFS of high-intermediate and high-risk DLBCL, regardless of the cell of origin. This observation should be confirmed in a larger study.

Circulating endothelial cells are increased in chronic myeloid leukemia blast crisis

We measured circulating endothelial precursor cells (EPCs), activated circulating endothelial cells (aCECs), and mature circulating endothelial cells (mCECs) using four-color multiparametric flow cytometry in the peripheral blood of 84 chronic myeloid leukemia (CML) patients and 65 healthy controls; and vascular endothelial growth factor (VEGF) by quantitative real-time PCR in 50 CML patients and 32 healthy controls. Because of an increase in mCECs, the median percentage of CECs in CML blast crisis (0.0146%) was significantly higher than in healthy subjects (0.0059%, P<0.01) and in the accelerated phase (0.0059%, P=0.01). There were no significant differences in the percentages of CECs in chronic- or active-phase patients and healthy subjects (P>0.05). In addition, VEGF gene expression was significantly higher in all phases of CML: 0.245 in blast crisis, 0.320 in the active phase, and 0.330 in chronic phase patients than it was in healthy subjects (0.145). In conclusion, CML in blast crisis had increased levels of CECs and VEGF gene expression, which may serve as markers of disease progression and may become targets for the management of CML.

Possible benefit of consolidation therapy with high-dose cytarabine on overall survival of adults with non-promyelocytic acute myeloid leukemia

In adults with non-promyelocytic acute myeloid leukemia (AML), high-dose cytarabine consolidation therapy has been shown to influence survival in selected patients, although the appropriate doses and schemes have not been defined. We evaluated survival after calculating the actual dose of cytarabine that patients received for consolidation therapy and divided them into 3 groups according to dose. We conducted a single-center, retrospective study involving 311 non-promyelocytic AML patients with a median age of 36 years (16-79 years) who received curative treatment between 1978 and 2007. The 131 patients who received cytarabine consolidation were assigned to study groups by their cytarabine dose protocol. Group 1 (n=69) received <1.5 g/m2 every 12 h on 3 alternate days for up to 4 cycles. The remaining patients received high-dose cytarabine (≥1.5 g/m2 every 12 h on 3 alternate days for up to 4 cycles). The actual dose received during the entire consolidation period in these patients was calculated, allowing us to divide these patients into 2 additional groups. Group 2 (n=27) received an intermediate-high-dose (<27 g/m2), and group 3 (n=35) received a very-high-dose (≥27 g/m2). Among the 311 patients receiving curative treatment, the 5-year survival rate was 20.2% (63 patients). The cytarabine consolidation dose was an independent determinant of survival in multivariate analysis; age, karyotype, induction protocol, French-American-British classification, and de novo leukemia were not. Comparisons showed that the risk of death was higher in the intermediate-high-dose group 2 (hazard ratio [HR]=4.51; 95% confidence interval [CI]: 1.81-11.21) and the low-dose group 1 (HR=4.43; 95% CI: 1.97-9.96) than in the very-high-dose group 3, with no significant difference between those two groups. Our findings indicated that very-high-dose cytarabine during consolidation in adults with non-promyelocytic AML may improve survival.

É necessária a realização de biópsia de medula óssea bilateral para o estadiamento do linfoma difuso de grandes células B

This retrospective study aims to analyze the usefulness of bilateral bone marrow biopsy in bone marrow infiltration by diffuse large B-cell lymphoma (DLBCL). Our objectives were to assess the incidence of unilateral BM involvement by DLBCL and compare fragment length obtained from positive and negative samples for infiltration. Furthermore, we compared the differences between unilateral and bilateral infiltration correlating with lactic dehydrogenase (LDH) and computerized tomography (CT) staging. We evaluated 268 cases of DLBCL and observed medullary infiltration in 34 cases (13%). It was not possible to evaluate 6 out of 34 cases. 70 BM fragments were reviewed as to the presence or absence of infiltration and length. The mean number of fragments per case was 2.5; the mean BM fragment length was 11.01 mm (± 5.12 mm) and the mean BM fragment length per case was 27.53 mm. There was unilateral BM infiltration in six cases (21.4%). There were no differences in the mean fragment length as to the presence/absence of infiltration 10.95 mm (± 5.2 mm) versus 11.57 mm, p > 0.05, respectively. There were no differences in 23 cases between the comparison of unilateral medullary infiltration versus bilateral with lactic dehydrogenase and CT staging. We concluded that bilateral bone marrow biopsy was superior to unilateral because it may increase by 21.4% the detection of BM involvement by DLBCL.

Significado prognóstico dos graus histológicos do linfoma de Hodgkin do tipo esclerose nodular

INTRODUCAO: A esclerose nodular (EN), do tipo histologico frequente do linfoma de Hodgkin (LH), apresenta grande variabilidade em sua composicao celular. Na decada de 80, pesquisadores do British National Lymphoma Investigation (BNLI) propuseram uma subclassificacao histologica do LH EN. Eles identificaram dois graus histologicos - o LH EN grau I (LH EN I) e o LH EN grau II (LH EN II) - e demonstraram que os portadores de LH EN II apresentavam menor sobrevida em comparacao aos portadores de LH EN I. Outros estudos, entretanto, nao reproduziram esses achados. OBJETIVO: Avaliar o significado prognostico da graduacao histologica proposta pelo BNLI. MATERIAIS E METODOS: Estudo retrospectivo que incluiu 69 casos de LH EN. Mais de 90% dos casos foram tratados com terapia combinada ou quimioterapia exclusiva, nao havendo diferenca no tipo de tratamento oferecido a depender do grau histologico. RESULTADOS: Trinta e cinco casos (51%) foram classificados com EN I e 34 (49%) como EN II. Nao observamos diferencas na distribuicao de outros fatores prognosticos entre os portadores dos dois graus. Remissao completa apos o tratamento inicial foi obtida em 85,7% dos casos de EN I e em 82,4% dos casos de EN II (p = 0,75). A probabilidade estimada de sobrevida global em cinco anos foi de 67% para EN I e de 83,5% para EN II (p = 0,13) e a taxa de sobrevida livre de doenca em cinco anos foi de 85,2% versus 87%, respectivamente (p = 0,72). CONCLUSAO: Nesta populacao de pacientes uniformemente tratados a graduacao histologica BNLI nao esteve associada com o prognostico do LH EN.