Internal tandem duplication of the FLT3 gene confers poor overall survival in patients with acute promyelocytic leukemia treated with all-trans retinoic acid and anthracycline-based chemotherapy: an International Consortium on Acute Promyelocytic Leukemia study
Lucena-Araujo A.R.,Center for Cell Based Therapy |
Kim H.T.,Dana-Farber Cancer Institute |
Jacomo R.H.,Center for Cell Based Therapy |
Melo R.A.,Fundacao HEMOPE |
And 24 more authors.
Annals of Hematology | Year: 2014
Activating internal tandem duplication (ITD) mutations in the fms-like tyrosine kinase 3 (FLT3) gene (FLT3-ITD) are associated with poor outcome in acute myeloid leukemia, but their prognostic impact in acute promyelocytic leukemia (APL) remains controversial. Here, we screened for FLT3-ITD mutations in 171 APL patients, treated with all-trans retinoic acid (ATRA) and anthracycline-based chemotherapy. We identified FLT3-ITD mutations in 35 patients (20 %). FLT3-ITD mutations were associated with higher white blood cell counts (P < 0.0001), relapse-risk score (P = 0.0007), higher hemoglobin levels (P = 0.0004), higher frequency of the microgranular morphology (M3v) subtype (P = 0.03), and the short PML/RARA (BCR3) isoform (P < 0.0001). After a median follow-up of 38 months, FLT3-ITDpositive patients had a lower 3-year overall survival rate (62 %) compared with FLT3-ITDnegative patients (82 %) (P = 0.006). The prognostic impact of FLT3-ITD on survival was retained in multivariable analysis (hazard ratio: 2.39, 95 % confidence interval [CI] 1.17–4.89; P = 0.017). Nevertheless, complete remission (P = 0.07), disease-free survival (P = 0.24), and the cumulative incidence of relapse (P = 0.94) rates were not significantly different between groups. We can conclude that FLT3-ITD mutations are associated with several hematologic features in APL, in particular with high white blood cell counts. In addition, FLT3-ITD may independently predict a shorter survival in patients with APL treated with ATRA and anthracycline-based chemotherapy. © 2014, Springer-Verlag Berlin Heidelberg.
Berzoti-Coelho M.G.,University of Sao Paulo |
Ferreira A.F.,University of Sao Paulo |
Ferreira A.F.,Center for Cell Based Therapy |
de Souza Nunes N.,University of Sao Paulo |
And 16 more authors.
Blood Cells, Molecules, and Diseases | Year: 2016
Chronic Myeloid Leukemia (CML), Polycythemia Vera (PV), Essential Thrombocythemia (ET) and Primary Myelofibrosis (PMF) are Myeloproliferative Neoplasms (MPN) characterized by clonal myeloproliferation without cell maturation impairment. CML pathogenesis is associated with the Ph chromosome leading to BCR-ABL tyrosine-kinase constitutive expression. The Ph negative MPN (PV, ET and PMF) are characterized by the mutation JAK2V617F of the JAK2 protein in the auto-inhibitory JH2 domain, which is found in most PV patients and in approximately half of ET and PMF patients. Considerable effort is being made to understand the role of JAK2V617F at the MPN initiation and to clarify the pathogenesis and apoptosis resistance in CML, PV, ET and PMF patients. In the present investigation, we evaluated the Death Inducer-Obliterator (DIDO) (variants DIDO 1, 2 and 3) levels in CML, PV, ET and PMF patients. Our data reported the DIDO 1, 2 and 3 differential expressions in Myeloproliferative Neoplasms. © 2016 Elsevier Inc.
de Araujo L.F.,University of Sao Paulo |
de Araujo L.F.,Center for Cell Based Therapy |
Fonseca A.S.,University of Sao Paulo |
Fonseca A.S.,Center for Cell Based Therapy |
And 16 more authors.
Tumor Biology | Year: 2015
Mitochondrial dysfunction is regarded as a hallmark of cancer progression. In the current study, we evaluated mitochondrial genome instability and copy number in colorectal cancer using Next Generation Sequencing approach and qPCR, respectively. The results revealed higher levels of heteroplasmy and depletion of the relative mtDNA copy number in colorectal adenocarcinoma. Adenocarcinoma samples also presented an increased number of mutations in nuclear genes encoding proteins which functions are related with mitochondria fusion, fission and localization. Moreover, we found a set of mitochondrial and nuclear genes, which cooperate in the same mitochondrial function simultaneously mutated in adenocarcinoma. In summary, these results support an important role for mitochondrial function and genomic instability in colorectal tumorigenesis. © 2015, International Society of Oncology and BioMarkers (ISOBM).
Sangiorgi B.,University of Sao Paulo |
Sangiorgi B.,Center for Cell Based Therapy |
Panepucci R.A.,University of Sao Paulo |
Panepucci R.A.,Center for Cell Based Therapy
Stem Cells International | Year: 2016
In the last decade, the immunomodulatory properties of mesenchymal stromal cells (MSCs) have attracted a lot of attention, due to their potential applicability in the treatment of graft-versus-host disease (GVHD), a condition frequently associated with opportunistic infections. The present review addresses how Pathogen-Associated Molecular Patterns (PAMPS) modulate the immunosuppressive phenotype of human MSCs by signaling through Toll-like receptors (TLRs). Overall, we observed that regardless of the source tissue, human MSCs express TLR2, TLR3, TLR4, and TLR9. Stimulation of distinct TLRs on MSCs elicits distinct inflammatory signaling pathways, differentially influencing the expression of inflammatory factors and the ability of MSCs to suppress the proliferation of immune system cells. The capacity to enhance the immunosuppressive phenotype of MSCs through TLRs stimulation might be properly elucidated in order to improve the MSC-based immunotherapy against GVHD. © 2016 Bruno Sangiorgi and Rodrigo Alexandre Panepucci.
Figueiredo L.M.,University of Sao Paulo |
Figueiredo L.M.,Center for Cell based Therapy |
Costa E.B.O.,University of Sao Paulo |
Costa E.B.O.,Center for Cell based Therapy |
And 6 more authors.
Cellular Reprogramming | Year: 2015
Hematopoietic cells (HCs) and endothelial cells (ECs) can be produced in vitro from human embryonic stem cells (hESCs), but the differentiation systems used are still inefficient. To overcome this obstacle, it is necessary to understand the differentiation process. One of the methods used to obtain HCs and ECs from hESCs is their co-culture with stromal cells. The soluble factors secreted by these cells and cell-cell contact have a great impact on the differentiation process. Here, we performed comparative proteomic analyses of proteins obtained from the total extract of OP9 stromal cells and secreted by these cells before and during in vitro generation of HCs and ECs (hematoendothelial) from hESCs. We identified a total of 83 secreted and 759 intracellular proteins during differentiation. Twenty-five secreted and 181 proteins from the total extract were more abundant. Some secreted proteins are involved in cell-matrix interactions and HC and/or EC development. Moreover, 13 proteins of the total extract from OP9 cells that were exclusive/or more abundant during differentiation are involved in the Nrf2/Nfe2l2 gene pathway, that is, they are described to have a key role in oxidative stress and in hematopoietic development and maturation. Our proteomic profiles provide valuable insight about the proteins involved in in vitro hematoendothelial cell generation and in the future they might be used to optimize the differentiation process and produce both cell types in vitro. © Mary Ann Liebert, Inc. 2015.
Martins C.S.,University of Sao Paulo |
Santana-Lemos B.A.,University of Sao Paulo |
Santana-Lemos B.A.,Center for Cell based Therapy |
Saggioro F.P.,University of Sao Paulo |
And 6 more authors.
Journal of Endocrinological Investigation | Year: 2015
Purpose: Telomere dysfunction and telomerase activation underlie cancer transformation. This study aims to investigate the contribution of telomere biology to pituitary tumor behavior. Subjects and methods: Samples from 50 patients with pituitary tumors (11 ACTH-secreting, 18 GH-secreting, and 21 non-secreting tumors) and 7 subjects without pituitary lesions were collected. The expressions of telomerase essential components TERT and TERC and tumor telomere content were measured by quantitative PCR techniques. Results: Telomerase (TERT) expression was detected in 36 % of tumors. No correlation was observed between TERT and TERC expression level and tumor size in any tumor type. There was no association between gene expression and clinical findings. Telomere content (T/S ratio) was similar between pituitary adenomas (0.39 ± 0.16) and normal pituitaries (0.47 ± 0.12; p = 0.24) and also was between the different adenoma types: ACTH-secreting (0.43 ± 0.08), GH-secreting (0.31 ± 0.12), and non-secreting (0.42 ± 0.20; p = 0.10) tumors. Conclusions: The telomere content and expression of telomerase components are comparable between normal pituitary glands and tumor tissues, suggesting that telomere biology does not play an important role in pituitary tumor development. © 2015 Italian Society of Endocrinology (SIE).
Poersch A.,University of Sao Paulo |
Poersch A.,Center for Cell Based Therapy |
Grassi M.L.,University of Sao Paulo |
Grassi M.L.,Center for Cell Based Therapy |
And 12 more authors.
Journal of Proteomics | Year: 2016
Tumor fluid samples have emerged as a rich source for the identification of ovarian cancer in the context of proteomics studies. To uncover differences among benign and malignant ovarian samples, we performed a quantitative proteomic study consisting of albumin immunodepletion, isotope labeling with acrylamide and in-depth proteomic profiling by LC-MS/MS in a pool of 10 samples of each histological type. 1135 proteins were identified, corresponding to 505 gene products. 223 proteins presented associated quantification and the comparative analysis of histological types revealed 75 differentially abundant proteins. Based on this, we developed a panel for targeted proteomic analysis using the multiple reaction monitoring (MRM) method for validation of 51 proteins in individual samples of high-grade serous ovarian tumor fluids (malignant) and benign serous cystadenoma tumor fluids. This analysis showed concordant results in terms of average amounts of proteins, and APOE, SERPINF2, SERPING1, ADAM17, CD44 and OVGP1 were statistically significant between benign and malignant group. The results observed in the MRM for APOE were confirmed by western blotting, where APOE was more abundant in malignant samples. This molecular signature can contribute to improve tumor stratification and shall be investigated in combination with current biomarkers in larger cohorts to improve ovarian cancer diagnosis. Biological significance Despite advances in cancer research, ovarian cancer has a high mortality and remains a major challenge due to a number of particularities of the disease, especially late diagnosis caused by vague clinical symptoms, the cellular and molecular heterogeneity of tumors, and the lack of effective treatment. Thus, efforts are directed to better understand this neoplasia, its origin, development and, particularly the identification and validation of biomarkers for early detection of the disease in asymptomatic stage. In the present work, we confirmed by MRM method in individual ovarian tumor fluid samples the regulation of 27 proteins out of 33 identified in a highthroughput study. We speculate that the presence and/or differential abundance observed in tumor fluid is a cooperation primarily of high rates of secretion of such tumor proteins to extra tumor environment that will at the end accumulate in plasma, and also the accumulation of acute-phase proteins throughout the entire body. On top of that, consideration of physiological influences in the interpretation of expression observed, including age, menopause status, route-of-elimination kinetics and metabolism of the tumor marker, coexisting disease, hormonal imbalances, life-style influences (smoking, alcoholism, obesity), among others, are mandatory to enable the selection of good protein tumor marker candidates for extensive validation. © 2016 Elsevier B.V.
Cle D.V.,University of Sao Paulo |
Cle D.V.,Center for Cell Based Therapy |
Santana-Lemos B.,University of Sao Paulo |
Santana-Lemos B.,Center for Cell Based Therapy |
And 7 more authors.
Cytotherapy | Year: 2015
Background aims: For patients with aplastic anemia (AA) who are refractory to anti-thymocyte globulin (ATG) and cyclosporine, a second course of immunosuppression is successful in only one-fourth to one-third of cases. Methods: We conducted a phase 1/2 study to evaluate the addition of two to five weekly intravenous infusions of allogeneic unrelated non-human leukocyte antigen-matched bone marrow-derived mesenchymal stromal cells (MSCs) (median, 2.7 × 106 cells/kg/infusion; range, 1.3-4.5) to standard rabbit ATG and cyclosporine in nine patients with refractory or relapsed AA. Results: After a median follow-up of 20 months, no infusion-related adverse event was observed, but four deaths occurred as the result of heart failure and bacterial or invasive fungal infections; only two patients achieved partial hematologic responses at 6 months. We failed to demonstrate by fluorescence in situ hybridization or variable number tandem repeat any MSC engraftment in patient marrow 30, 90 or 180 days after infusions. Conclusions: Infusion of allogeneic MSCs in AA is safe but does not improve clinical hematologic response or engraft in recipient bone marrow. This study was registered at clinicaltrials.gov, identifier: NCT01297972. © 2015 International Society for Cellular Therapy.
Gutierrez-Rodrigues F.,University of Ribeirão Preto |
Gutierrez-Rodrigues F.,Center for Cell based Therapy |
Santana-Lemos B.A.,University of Ribeirão Preto |
Santana-Lemos B.A.,Center for Cell based Therapy |
And 6 more authors.
PLoS ONE | Year: 2014
Telomere length measurement is an essential test for the diagnosis of telomeropathies, which are caused by excessive telomere erosion. Commonly used methods are terminal restriction fragment (TRF) analysis by Southern blot, fluorescence in situ hybridization coupled with flow cytometry (flow-FISH), and quantitative PCR (qPCR). Although these methods have been used in the clinic, they have not been comprehensively compared. Here, we directly compared the performance of flow-FISH and qPCR to measure leukocytes' telomere length of healthy individuals and patients evaluated for telomeropathies, using TRF as standard. TRF and flow-FISH showed good agreement and correlation in the analysis of healthy subjects (R2 = 0.60; p<0.0001) and patients (R2 = 0.51; p<0.0001). In contrast, the comparison between TRF and qPCR yielded modest correlation for the analysis of samples of healthy individuals (R2 = 0.35; p<0.0001) and low correlation for patients (R2 = 0.20; p = 0.001); Bland-Altman analysis showed poor agreement between the two methods for both patients and controls. Quantitative PCR and flow-FISH modestly correlated in the analysis of healthy individuals (R2 = 0.33; p<0.0001) and did not correlate in the comparison of patients' samples (R2 = 0.1, p = 0.08). Intra-assay coefficient of variation (CV) was similar for flow-FISH (10.8±7.1%) and qPCR (9.5±7.4%; p = 0.35), but the inter-assay CV was lower for flow-FISH (9.6±7.6% vs. 16±19.5%; p = 0.02). Bland-Altman analysis indicated that flow-FISH was more precise and reproducible than qPCR. Flow-FISH and qPCR were sensitive (both 100%) and specific (93% and 89%, respectively) to distinguish very short telomeres. However, qPCR sensitivity (40%) and specificity (63%) to detect telomeres below the tenth percentile were lower compared to flow-FISH (80% sensitivity and 85% specificity). In the clinical setting, flow-FISH was more accurate, reproducible, sensitive, and specific in the measurement of human leukocyte's telomere length in comparison to qPCR. In conclusion, flow-FISH appears to be a more appropriate method for diagnostic purposes. Copyright: © 2014 Gutierrez-Rodrigues et al.