Maria Aparecida Zanichelli

Epidemiology of bacteremia and factors associated with multi-drug- resistant gram-negative bacteremia in hematopoietic stem cell transplant recipients

The incidence of Gram-negative bacteremia has increased in hematopoietic stem cell transplant (HSCT) recipients. We prospectively collected data from 13 Brazilian HSCT centers to characterize the epidemiology of bacteremia occurring early post transplant, and to identify factors associated with infection due to multi-drug-resistant (MDR) Gram-negative isolates. MDR was defined as an isolate with resistance to at least two of the following: third- or fourth-generation cephalosporins, carbapenems or piperacillin-tazobactam. Among 411 HSCT, fever occurred in 333, and 91 developed bacteremia (118 isolates): 47% owing to Gram-positive, 37% owing to Gram-negative, and 16% caused by Gram-positive and Gram-negative bacteria. Pseudomonas aeruginosa (22%), Klebsiella pneumoniae (19%) and Escherichia coli (17%) accounted for the majority of Gram-negative isolates, and 37% were MDR. These isolates were recovered from 20 patients, representing 5% of all 411 HSCT and 22% of the episodes with bacteremia. By multivariate analysis, treatment with third-generation cephalosporins (odds ratio (OR) 10.65, 95% confidence interval (CI) 3.75–30.27) and being at one of the hospitals (OR 9.47, 95% CI 2.60–34.40) were associated with infection due to MDR Gram-negative isolates. These findings may have important clinical implications in the decision of giving prophylaxis and selecting the empiric antibiotic regimen.

ABCB1 haplotype is associated with major molecular response in chronic myeloid leukemia patients treated with standard-dose of imatinib

BACKGROUND Imatinib mesylate (IM) is a selective tyrosine kinase inhibitor used for treating chronic myeloid leukemia (CML). IM has high efficacy, however some individuals develop a resistance due to impaired bioavailability. Polymorphisms in genes encoding membrane transporters such as ABCB1 have been associated with differences in protein expression and function that influence the response to several drugs. AIM To investigate the relationship of ABCB1 polymorphisms with markers of response to IM in patients with CML. METHODS One hundred eighteen CML patients initially treated with a standard dose of IM (400mg/day) for 18months were selected at two health centers in Sao Paulo City, Brazil. The response criteria were based on the European LeukemiaNet recommendations. ABCB1 polymorphisms c.1236C>T (rs1128503), c.3435C>T (rs1045642) and c.2677G>T/A (rs2032582) were evaluated by PCR-RFLP. RESULTS ABCB1 polymorphisms were not related with a risk for CML in this sample population (p<0.05). In the CML group, frequencies of ABCB1 SNPs were similar between responder and non-responder patients (p>0.05). In the responder group, the frequency of ABCB11236CT/2677GT/3435CT haplotype was higher in patients with major molecular response (MMR) (51.7%) than in patients without MMR (8.3%, p=0.010). Furthermore, carriers of this haplotype had increased the probability of reaching the MMR compared with the non-carriers (OR: 11.8; 95% CI: 1.43-97.3, p=0.022). CONCLUSIONS The ABCB1 1236CT/2677GT/3435CT haplotype is positively associated with the major molecular response to IM in CML patients.

BCR-ABL-mediated upregulation of PRAME is responsible for knocking down TRAIL in CML patients.

Tumor necrosis factor-related apoptosis-inducing ligand—TNFSF10 (TRAIL), a member of the TNF-α family and a death receptor ligand, was shown to selectively kill tumor cells. Not surprisingly, TRAIL is downregulated in a variety of tumor cells, including BCR–ABL-positive leukemia. Although we know much about the molecular basis of TRAIL-mediated cell killing, the mechanism responsible for TRAIL inhibition in tumors remains elusive because (a) TRAIL can be regulated by retinoic acid (RA); (b) the tumor antigen preferentially expressed antigen of melanoma (PRAME) was shown to inhibit transcription of RA receptor target genes through the polycomb protein, enhancer of zeste homolog 2 (EZH2); and (c) we have found that TRAIL is inversely correlated with BCR–ABL in chronic myeloid leukemia (CML) patients. Thus, we decided to investigate the association of PRAME, EZH2 and TRAIL in BCR–ABL-positive leukemia. Here, we demonstrate that PRAME, but not EZH2, is upregulated in BCR–ABL cells and is associated with the progression of disease in CML patients. There is a positive correlation between PRAME and BCR–ABL and an inverse correlation between PRAME and TRAIL in these patients. Importantly, knocking down PRAME or EZH2 by RNA interference in a BCR–ABL-positive cell line restores TRAIL expression. Moreover, there is an enrichment of EZH2 binding on the promoter region of TRAIL in a CML cell line. This binding is lost after PRAME knockdown. Finally, knocking down PRAME or EZH2, and consequently induction of TRAIL expression, enhances Imatinib sensibility. Taken together, our data reveal a novel regulatory mechanism responsible for lowering TRAIL expression and provide the basis of alternative targets for combined therapeutic strategies for CML.

Relationship between SLCO1B3 and ABCA3 polymorphisms and imatinib response in chronic myeloid leukemia patients

Abstract Background Genetic variations in membrane transporters may contribute to imatinib mesylate (IM) resistance in chronic myeloid leukemia (CML). Objective To investigate the relationship between SLCO1B3, SLCO1A2, and ABCA3 polymorphisms and IM response in CML patients. Methods Patients in chronic phase CML (N = 118) were studied. All patients were treated with a standard dose of IM (400 mg/day) and classified into one of the two groups according to their responses. Major molecular response (MMR) and complete molecular response (CMR) were evaluated. Criteria for response failure were established according to European LeukemiaNet (2009). Analysis of the SLCO1B3 c.334T > G (rs4149117) and c.699G > A (rs7311358), SLCO1A2 c.516A > C (rs11568563) and c.-62-361G > A (rs3764043), and ABCA3 c.1755C > G (rs323043) and c.4548-191C > A (rs150929) polymorphisms was carried out by real-time polymerase chain reaction. Results SLCO1A2 and ABCA3 polymorphisms have similar frequencies between responders and non-responders. SLCO1B3 699GG and 344TT genotypes were more frequent in the responder group (63.8%) than in the non-responder group (44.7%, P = 0.042). Furthermore, carriers of 699GA/AA and 334TG/GG genotypes presented a higher probability of not responding to the standard dose of IM (odds ratio: 2.17; 95% confidence interval: 1.02–4.64, P = 0.04). Poor CMR for ABCA3 4548-91C > A was observed in patients with the CC/CA genotype when compared to AA carriers in the responder group (P = 0.014). Conclusions SLCO1B3 699GG and 344TT genotypes are associated with non-response to IM, while ABCA3 4548-91 CC/CA genotypes are related to poor CMR in CML patients treated with standard-dose imatinib.

Retrospective epidemiological study of Latin American patients with transfusional hemosiderosis: the first Latin American epidemiological study in iron overload – the RELATH study

Abstract The retrospective epidemiological study of Latin Americans with transfusional hemosiderosis is the first regional patient registry to gather data regarding the burden of transfusional hemosiderosis and patterns of care in these patients. Retrospective and cross-sectional data were collected on patients ⩾2 years with selected chronic anemias and minimum 20 transfusions. In the 960 patients analyzed, sickle-cell disease (48·3%) and thalassemias (24·0%) were the most frequent underlying diagnoses. The registry enrolled 355 pediatric patients (187 with sickle-cell disease/94 with thalassemia). Serum ferritin was the most frequent method used to detect iron overload. Complications from transfusional hemosiderosis were reported in ∼80% of patients; hepatic (65·3%), endocrine (27·5%), and cardiac (18·2%) being the most frequent. These data indicate that hemoglobinopathies and complications due to transfusional hemosiderosis are a significant clinical problem in the Latin American population with iron overload. Chelation therapy is used insufficiently and has a high rate of discontinuation.

Apoptosis-related gene expression profile in chronic myeloid leukemia patients after imatinib mesylate and dasatinib therapy

Background/Aims: We investigated the effects of tyrosine kinase inhibitors (TKIs) on the expression of apoptosis-related genes (BCL-2 and death receptor family members) in chronic myeloid leukemia (CML) patients. Methods: Peripheral blood mononuclear cells from 32 healthy subjects and 26 CML patients were evaluated before and after treatment with imatinib mesylate (IM) and dasatinib (DAS) by quantitative PCR. Results: Anti-apoptotic genes (c-FLIP and MCL-1) were overexpressed and the pro-apoptotic BIK was reduced in CML patients. Expression of BMF, A1, c-FLIP, MCL-1, CIAP-2 and CIAP-1 was modulated by DAS. In IM-resistant patients, expression of A1, c-FLIP, CIAP-1 and MCL-1 was upregulated, and BCL-2, CIAP-2, BAK, BAX, BIK and FASL expression was downregulated. Conclusion: Taken together, our results point out that, in CML, DAS interferes with the apoptotic machinery regulation. In addition, the data suggest that apoptosis-related gene expression profiles are associated with primary resistance to IM.

Estudo retrospectivo do tratamento de leucemia mielóide aguda com o transplante de medula óssea: a experiência brasileira

Data from the International Bone Marrow Transplant Registry (IBMTR) contribute for the improvement of Bone Marrow Transplant (BMT) worldwide. We studied the Brazilian experience in BMT for AML to compare this with international data. We performed a retrospective study by sending questionnaires to 16 BMT centers regarding clinical and treatment variables. Statistical analyses concerning autologous BMT (autoBMT) and allogeneic BMT (alloBMT) were performed using the Kaplan-Meier method and the log-rank test. All p-values were two-tailed. We collected data from 731 patients (205 autoBMT and 526 alloBMT). Median overall survival (OS) for autoBMT patients was longer than alloBMT patients (1035 vs. 466 days, p=0.0012). AlloBMT stem cell source (SCS): 73% bone marrow stem cell (BMSC), 23% peripheral blood stem cells (PBSC) and 4% umbilical cord blood. Among the autoBMT patients, the SCS was 63% PBSC, 22% BMSC and 15% both. The SCS did not impact on OS. There was no difference in OS between different FAB classifications in the alloBMT group, but in the autoBMT the M3 patients had longer survival. As expected, the main cause of mortality among autoBMT patients was related to disease relapse (60%), while in the alloBMT, to infection (38%). In both groups we found longer OS in first complete remission (1CR) compared to second (2CR) and other (p<0.0001), and longer OS in de novo AML than in secondary. In the alloBMT group we found more patients with advanced disease (60%), while in the autoBMT group, we found more M3 patients (24%), which could explain the difference in OS. Most of our results are in accordance with IBMTR data. One should consider the fact that this is a retrospective study and our findings should be analysed with caution.

Indicações em transplante de células-tronco hematopoéticas em pacientes adultos com leucemia linfoide aguda

In acute lymphoblastic leukemia, accumulation and proliferation of immature cells infiltration characterise a heterogeneous entity, featuring a wide variety of clinical and biological aspects. In the adult LLA concentration of high-risk prognosis factors such as age, B-cell, chromosomic changes, and chiefly the presence of chromosome positive Ph. Considerations of high morbidity and mortality rates related to haematopoietic stem cell transplantation (TCTH) have generated controversy about this therapeutic modality in adult patients with LLA in first remission (1st RC). The results of conventional therapy with chemotherapy in contrast with different risk groups of patients with LLA, has been used for the indication of TCTH. Thus we present the algorithm indications of haematopoietic stem cell transplantation in adult patients with LLA. Rev.

Differential expression of apoptomiRs in myeloproliferative neoplasms.

MicroRNAs are small non-coding RNAs that inhibit posttranscriptional gene expression through deadenylation and cleavage. Th ese molecules play crucial roles in regulating cell diff erentiation, proliferation and apoptosis [1]. MicroRNA expression levels are strongly associated with numerous human pathogenic diseases [2]. Th us, the discovery of microRNAs has opened up a wide range of possibilities regarding the understanding of disease pathophysiology and the development of new therapeutic agents. Recent studies show that microRNAs may participate in the regulation of apoptosis machinery (apoptomiRs). Apoptosis is a programmed cell death, orchestrated by the BCL-2 protein family, death receptors and inhibitor of apoptosis proteins (IAPs) [3]. Deregulated apoptosis is associated with tumorigenesis and autoimmunity [3] and seems to contribute to myeloproliferative neoplasms (MPNs), which are clonal disorders characterized by myeloaccumulation and in most cases by the presence of a somatic mutation in Janus kinase 2 (JAK2) [4]. Our research group previously described abnormal apoptosis in patients with MPNs [5 – 7]. We found abnormal expression of death-receptor pathway-related genes such as FAIM and C-FLIP in leukocytes and CD34 � hematopoietic stem cells (HSCs) [5], and deregulation of BCL-2 family members associated with JAK2 mutation [6,7]. Th ese fi ndings led us to investigate the molecular mechanisms associated with apoptosis deregulation in patients with MPNs. In this study, we sought to identify microRNAs whose targets are apoptosis-related genes, investigate their expression in patients with MPNs, and correlate this with the expression of apoptosis-related genes and JAK2 mutation allele burden using three algorithms (websites: http://www.diana.pcbi. upenn.edu/cgi-bin/miRGen/v3/Targets.cgi; Microcosm Targets: http://www.ebi.ac.uk/enright-srv/microcosm/cgi-bin/ targets/v5/hit_list.pl?genome_id � 2964; and Target Scan: http://www.targetscan.org). Th e microRNAs and their respective target genes (apoptosis-related genes) predicted by the bioinformatics tools are as follows: miR-15a: BIK and BCL-2; miR-16: BID, BIK, BCL-2 and BCL xl; miR-26a: C-FLIP, CIAP-2 and MCL-1; miR-130b: CIAP-2; miR-21: CIAP-2, FASL and BCL-2; miR-29c: CIAP-1 and MCL-1; and miR-let-7d: A1, BAX, FAS and FASL. In this study, we investigated apoptomiRs miR-26a, -130b, -21, -29c, -let-7d, -15a and -16. We quantifi ed their expression in bone marrow CD34 � HSCs and peripheral blood leukocytes in 27 patients with polycythemia vera (PV), 25 with essential thrombocythemia (ET) and 12 with primary myelofi brosis (PMF) (26 males and 38 females, mean age: 60.5 years). Th e bone marrow control group comprised 14 bone mar

Matrix metalloproteinase-9 is consistently expressed in Hodgkin/Reed-Sternberg cells and has no impact on survival in patients with Epstein–Barr virus (EBV)-related and non-related Hodgkin lymphoma in Brazil

Clinical and histological features of classical Hodgkin lymphoma (cHL) are primarily due to the effects of cytokines and enzymes produced by Hodgkin/Reed-Sternberg (HRS) cells and their surrounding inflammatory cells. In EBV-related cancers, the expression of viral latent membrane protein 1 correlates with an increased MMP9 expression. In this study, we evaluated the prognostic relevance of MMP9 expression and EBV status in HRS cells in patients with cHL in Brazil. We selected 97 patients with cHL for EBV and MMP9 detection. EBV was detected in 52.5%, and MMP9 expression positivity was found in 87.6%. Of all cases, there was no correlation between MMP9 expression and EBV status. Response to treatment and relapse rate was independent of MMP9 expression and EBV status. MMP9 positivity did not influence overall survival and event-free survival. The consistent and increased intensity of MMP9 expression in HRS cells make this enzyme a potential target for therapy.