Global Proteomics Analysis of Bone Marrow: Establishing Talin-1 and Centrosomal Protein of 55 kDa as Potential Molecular Signatures for Myelodysplastic Syndromes.

Myelodysplastic syndrome (MDS) is a hematological disorder characterized by abnormal stem cell differentiation and a high risk of acute myeloid leukemia transformation. Treatment options for MDS are still limited, making the identification of molecular signatures for MDS progression a vital task. Thus, we evaluated the proteome of bone marrow plasma from patients (n = 28) diagnosed with MDS with ring sideroblasts (MDS-RS) and MDS with blasts in the bone marrow (MDS-EB) using label-free mass spectrometry. This strategy allowed the identification of 1,194 proteins in the bone marrow plasma samples. Polyubiquitin-C (UBC), moesin (MSN), and Talin-1 (TLN1) showed the highest abundances in MDS-EB, and centrosomal protein of 55 kDa (CEP55) showed the highest relative abundance in the bone marrow plasma of MDS-RS patients. In a follow-up, in the second phase of the study, expressions of UBC, MSN, TLN1, and CEP55 genes were evaluated in bone marrow mononuclear cells from 45 patients by using qPCR. This second cohort included only seven patients from the first study. CEP55, MSN, and UBC expressions were similar in mononuclear cells from MDS-RS and MDS-EB individuals. However, TLN1 gene expression was greater in mononuclear cells from MDS-RS (p = 0.049) as compared to MDS-EB patients. Irrespective of the MDS subtype, CEP55 expression was higher (p = 0.045) in MDS patients with abnormal karyotypes, while MSN, UBC, and TALIN1 transcripts were similar in MDS with normal vs. abnormal karyotypes. In conclusion, proteomic and gene expression approaches brought evidence of altered TLN1 and CEP55 expressions in cellular and non-cellular bone marrow compartments of patients with low-risk (MDS-RS) and high-risk (MDS-EB) MDSs and with normal vs. abnormal karyotypes. As MDS is characterized by disrupted apoptosis and chromosomal alterations, leading to mitotic slippage, TLN1 and CEP55 represent potential markers for MDS prognosis and/or targeted therapy.

c 9253-6T > c REV3L: A novel marker of poor prognosis in Myelodysplastic syndrome

Myelodysplastic syndromes (MDS) are a heterogeneous group of hematological malignancies characterized by dysplasias, ineffective hematopoiesis and risk of acute myeloid leukemia transformation. Approximately 90% of MDS patients present mutations in genes involved in various cell signaling pathways. Specialized DNA polymerases, such as POLN, POLI, POLK, POLQ, POLH, POLL and REV3L, insert a nucleotide opposite replication-blocking DNA lesions in an error-prone manner and, in this way, sometimes can actively promote the generation of mutation. For the best of our knowledge, has not been described the mutations of these genes in MDS. DNA target sequencing CDS regions of the REV3L gene was performed in a 58-year-old man diagnosed as High Risk Myelodysplastic Syndrome. The patient presented very low hemoglobin, increased number of blasts, karyotype:47,XY,+8[6]/47,XY,del(7)(q32),+8[7], no response to hypomethylating therapy (decitabine), all markers of poor prognosis. Target sequencing identified a mutation c.9253-6T>C REV3L (Substitution - intronic) with VAF (variant allele frequency) = 16% considered pathogenic according to Functional Analysis through. Hidden Markov Models (FATHMM). This is the first evidence of REV3L mutation in MDS and, of utmost importance, associated with poor prognosis.

Immune Dysregulation and Recurring Mutations in Myelodysplastic Syndromes Pathogenesis.

Myelodysplastic syndromes (MDS) are clonal stem cell malignancies characterized by ineffective hematopoiesis leading to peripheral cytopenias and variable risk of progression to acute myeloid leukemia. Inflammation is associated with MDS pathogenesis. Several cytokines, reactive species of oxygen/nitrogen and growth factors are directly or indirectly involved in dysfunction of the MDS bone marrow (BM) microenvironment. Mutations in genes mainly regulating RNA splicing, DNA methylation and chromatin accessibility, transcription factors, signal transduction and the response to DNA damage contribute to ineffective hematopoiesis, genomic instability and MDS development. The inflammation-associated DNA damage in hematopoietic stem cells may also contribute to MDS development and progression with aggressive clinical characteristics. Many studies have aimed at clarifying mechanisms involved in the activity of immature myeloid cells as powerful modulators of the immune response and their correlation with aging, autoimmunity, and development of cancer. In this review, we explore recent advances and accumulating evidence uniting immune dysregulation, inflammaging and recurring mutations in the pathogenesis of MDS.

c.9253-6T>c REV3L: A novel marker of poor prognosis in Myelodysplastic syndrome.

Myelodysplastic syndromes (MDS) are a heterogeneous group of hematological malignancies characterized by dysplasias, ineffective hematopoiesis and risk of acute myeloid leukemia transformation. Approximately 90% of MDS patients present mutations in genes involved in various cell signaling pathways. Specialized DNA polymerases, such as POLN, POLI, POLK, POLQ, POLH, POLL and REV3L, insert a nucleotide opposite replication-blocking DNA lesions in an error-prone manner and, in this way, sometimes can actively promote the generation of mutation. For the best of our knowledge, has not been described the mutations of these genes in MDS. DNA target sequencing CDS regions of the REV3L gene was performed in a 58-year-old man diagnosed as High Risk Myelodysplastic Syndrome. The patient presented very low hemoglobin, increased number of blasts, karyotype:47,XY,+8[6]/47,XY,del(7)(q32),+8[7], no response to hypomethylating therapy (decitabine), all markers of poor prognosis. Target sequencing identified a mutation c.9253-6T>C REV3L (Substitution - intronic) with VAF (variant allele frequency)=16% considered pathogenic according to Functional Analysis through. Hidden Markov Models (FATHMM). This is the first evidence of REV3L mutation in MDS and, of utmost importance, associated with poor prognosis.

Differing clinical features between Japanese and Caucasian patients with myelodysplastic syndromes: Analysis from the International Working Group for Prognosis of MDS.

Clinical features of myelodysplastic syndromes (MDS) could be influenced by many factors, such as disease intrinsic factors (e.g., morphologic, cytogenetic, molecular), extrinsic factors (e.g, management, environment), and ethnicity. Several previous studies have suggested such differences between Asian and European/USA countries. In this study, to elucidate potential differences in primary untreated MDS between Japanese (JPN) and Caucasians (CAUC), we analyzed the data from a large international database collected by the International Working Group for Prognosis of MDS (300 and 5838 patients, respectively). JPN MDS were significantly younger with more severe cytopenias, and cytogenetic differences: less del(5q) and more +1/+1q, -1/del(1p), der(1;7), -9/del(9q), del(16q), and del(20q). Although differences in time to acute myeloid leukemia transformation did not occur, a significantly better survival in JPN was demonstrated, even after the adjustment for age and FAB subtypes, especially in lower, but not in higher prognostic risk categories. Certain clinical factors (cytopenias, blast percentage, cytogenetic risk) had different impact on survival and time to transformation to leukemia between the two groups. Although possible confounding events (e.g., environment, diet, and access to care) could not be excluded, our results indicated the existence of clinically relevant ethnic differences regarding survival in MDS between JPN and CAUC patients. The good performance of the IPSS-R in both CAUC and JP patients underlines that its common risk model is adequate for CAUC and JP.

Observational study of multiple myeloma in Latin America.

Relatively little is known about the outcomes of multiple myeloma in Latin America, a world region where incorporation of novel agents is generally slow. In the current retrospective-prospective study, we aimed to describe the patterns of care and treatment results in five Latin American countries. Between April 2007 and October 2009, patients who had been diagnosed from January 2005 to December 2007 were registered at 23 institutions from Argentina, Brazil, Chile, Mexico, and Peru. We divided patients into two cohorts, according to transplantation eligibility, and analyzed them with regard to first-line treatment and overall survival (OS). We analyzed a total of 852 patients, 46.9 % of whom were female. The median follow-up was 62 months. Among transplantation-ineligible patients (N = 461), the mean age was 67.4 years, approximately one third of patients received a thalidomide-based treatment in the first line, and the median OS was 43.0 months. Transplantation-eligible patients (N = 391) had a mean age of 54.7 years and a median OS of 73.6 months. Autologous transplantation was performed in 58.6 % of the patients for whom this procedure was initially planned and in only 26.9 % of the overall patients. Our long-term results reflect the contemporary literature for patients with multiple myeloma treated with autologous transplantation and thalidomide-based regimens in clinical trials and observational studies. However, further efforts are needed to approve and incorporate novel agents in Latin American countries, as well as to increase access to transplantation, in order to achieve the expected improvements in patient outcomes.

Time-dependent changes in mortality and transformation risk in MDS.

In myelodysplastic syndromes (MDSs), the evolution of risk for disease progression or death has not been systematically investigated despite being crucial for correct interpretation of prognostic risk scores. In a multicenter retrospective study, we described changes in risk over time, the consequences for basal prognostic scores, and their potential clinical implications. Major MDS prognostic risk scoring systems and their constituent individual predictors were analyzed in 7212 primary untreated MDS patients from the International Working Group for Prognosis in MDS database. Changes in risk of mortality and of leukemic transformation over time from diagnosis were described. Hazards regarding mortality and acute myeloid leukemia transformation diminished over time from diagnosis in higher-risk MDS patients, whereas they remained stable in lower-risk patients. After approximately 3.5 years, hazards in the separate risk groups became similar and were essentially equivalent after 5 years. This fact led to loss of prognostic power of different scoring systems considered, which was more pronounced for survival. Inclusion of age resulted in increased initial prognostic power for survival and less attenuation in hazards. If needed for practicability in clinical management, the differing development of risks suggested a reasonable division into lower- and higher-risk MDS based on the IPSS-R at a cutoff of 3.5 points. Our data regarding time-dependent performance of prognostic scores reflect the disparate change of risks in MDS subpopulations. Lower-risk patients at diagnosis remain lower risk whereas initially high-risk patients demonstrate decreasing risk over time. This change of risk should be considered in clinical decision making.

Myelodysplastic syndromes in South America: a multinational study of 1080 patients.

There are previously reported data describing differences between Asian and European patients with Myelodysplastic Syndromes (MDS), few direct comparisons based on cancer registration characteristics or using cohorts to validate scoring systems. This is the first study from South-America, which attempts to describe demographic, clinical features, and outcome of MDS patients. We retrospectively analyzed 1,080 patients with de novo MDS from Argentina (635), Brazil (345), and Chile (100). Chilean patients were younger (P = 0.001) with female preponderance (P = 0.071). Brazilian series showed a higher predominance of RARS subtype regarding FAB and WHO classifications (P < 0.001). Hemoglobin levels were significantly lower in Brazilian and Chilean series (P < 0.001), and Chilean series also showed a lower platelet count (P = 0.028), with no differences concerning the neutrophil count, % BM blast, and the distribution of cytogenetic risk groups (P > 0.05). Chilean series depicted a lower overall survival (OS; 35 months vs. 56 months-Argentine; 55 months-Brazil, P = 0.030), which was consistent with a higher predominance of the high-risk group according both to the IPSS and IPSS-R (P = 0.046 and P < 0.001). The IPSS-R system and its variables showed a good reproducibility to predict clinical outcome for the whole South-American population. Epidemiological and clinical characteristics, distribution among prognostic subgroups, the OS, and the access to disease modifying therapies were more similar between Argentinean and Brazilian compared with Chilean MDS series. This will need further analysis in a larger group of patients. Descriptive and comparative studies are necessary to establish epidemiological features useful for public health attitudes to generate suitable therapeutic schemes.

Revised international prognostic scoring system for myelodysplastic syndromes.

The International Prognostic Scoring System (IPSS) is an important standard for assessing prognosis of primary untreated adult patients with myelodysplastic syndromes (MDS). To refine the IPSS, MDS patient databases from international institutions were coalesced to assemble a much larger combined database (Revised-IPSS [IPSS-R], n = 7012, IPSS, n = 816) for analysis. Multiple statistically weighted clinical features were used to generate a prognostic categorization model. Bone marrow cytogenetics, marrow blast percentage, and cytopenias remained the basis of the new system. Novel components of the current analysis included: 5 rather than 3 cytogenetic prognostic subgroups with specific and new classifications of a number of less common cytogenetic subsets, splitting the low marrow blast percentage value, and depth of cytopenias. This model defined 5 rather than the 4 major prognostic categories that are present in the IPSS. Patient age, performance status, serum ferritin, and lactate dehydrogenase were significant additive features for survival but not for acute myeloid leukemia transformation. This system comprehensively integrated the numerous known clinical features into a method analyzing MDS patient prognosis more precisely than the initial IPSS. As such, this IPSS-R should prove beneficial for predicting the clinical outcomes of untreated MDS patients and aiding design and analysis of clinical trials in this disease.

Tissue doppler echocardiography detects preclinical markers of cardiac lesion in MDS patients.

Myelodysplastic syndrome (MDS) is a clonal hematopoietic stem cell disorder of elderly people. Cardiac dysfunction is a marker of grim prognosis in MDS. We evaluated cardiac dysfunction of MDS patients with or without transfusion dependency by tissue doppler echocardiography. We found the average values of ventricular end-systolic and end-diastolic volumes in transfusion dependency MDS group higher than others. These results were strongly correlated to hemoglobin levels. Tissue Doppler Echocardiography should be routinely performed in MDS patients to detect preclinical cardiac alterations and prevent more heart insults in this group of chronic anemic aged patients.

Síndromes mielodisplásicas e mielodisplásicas/mieloproliferativas / Myelodysplastic syndromes and diseases with myelodysplastic and myeloproliferative features

As síndromes mielodisplásicas (SMD) representam um grupo heterogêneo de doenças hematológicas caracterizadas por hematopoese ineficaz e risco aumentado de evolução para leucemia mieloide aguda. Neste artigo educativo são apresentados aspectos gerais da sua fisiopatologia, diagnóstico, apresentação histopatológica e seu papel no diagnóstico diferencial, classificação e estratificação prognóstica. Ressalta-se a importância da avaliação clínica e laboratorial, que inclui avaliação do sangue periférico e medula óssea: morfologia - aspirado medular e biópsia óssea -, citogenética, imunofenotipagem, além de dados evolutivos. O diagnóstico definitivo, em especial nos casos de baixo risco, deve considerar a exclusão de causas não clonais que podem, através de alterações dismielopoéticas reativas, simular a mielodisplasia, tais como infecções virais, principalmente pelo HIV. A nova classificação revisada da Organização Mundial da Saúde (OMS-2008) é apresentada e discutida.

Myelodysplastic syndromes (MDS) represent a heterogeneous group of hematologic disorders characterized by ineffective hematopoiesis and an increased risk of developing acute myeloid leukemia. In this educational article the general aspects of the physiopathology, diagnosis, and histopathological features of MDS and their role in differential diagnosis, classification and prognostic categorization are presented. The importance of clinical and laboratory evaluations, including peripheral blood and bone marrow analyses, including morphology - aspirate and core biopsy, cytogenetics, immunophenotype and careful serial follow-up is emphasized. Definite diagnosis of MDS, especially in low-risk subtypes, should consider the exclusion of disorders with reactive bone marrow alterations, such as viral infections for example HIV. The new revised World Health Organization (WHO-2008) classification is presented and discussed.

Management of transfusional iron overload in Latin America: current outlook and expert panel recommendations.

The results of a meeting of physicians convening in Latin America to develop expert opinions on the diagnosis, monitoring and treatment of iron overload are as follows. An accurate diagnosis can be obtained by neonatal screening for haemoglobinopathies, especially sickle cell disease and the thalassaemias. Disease-specific registries are needed to demonstrate the extent of the problem to health authorities. Disparities in the quantity and quality of blood products must be addressed, and uniform transfusion guidelines are necessary. Serum ferritin level is a feasible marker for iron overload in the region, while magnetic resonance imaging assessment can improve the diagnosis and monitoring of cardiac and liver iron content. Medical specialists, including radiologists, pathologists and others, and health authorities, can help to implement these methods and provide adequate resources. The recently available oral deferasirox can be used to conveniently administer iron chelation to transfusional iron-overloaded patients.

Efeito da bomba de infusão de soluções sobre o grau de hemólise em concentrados de hemácias / Effect of the infusion pumps of solutions on the degree of hemolysis

A lise das hemácias, de causa imune ou não imune, pode provocar a elevação da hemoglobina plasmática induzindo efeitos deletérios principalmente nos rins e no sistema cardiovascular. Este trabalho objetiva evidenciar o possível efeito hemolítico induzido pelo trauma mecânico provocado pela passagem das hemácias em diferentes modelos de bombas de infusão de soluções, em concentrados de hemácias com até dez dias de armazenamento, preparados com Sag-Manitol. Foram utilizados três modelos de bombas (Nutrimat, Infusomat Compact e Volumed), com quatro aparelhos de cada modelo, testados em três velocidades de infusão (120 mL/h, 240 mL/h e 360 mL/h). Os parâmetros utilizados para evidenciar o grau de hemólise foram o percentual de hemólise e os níveis de hemoglobina e potássio livres no plasma. As amostras foram coletadas antes da passagem do concentrado de hemácias pelas bombas, na metade do tempo de infusão e no final da infusão, nas diferentes velocidades. Não ocorreu variação significativa entre os valores dos parâmetros analisados nas amostras controle e naquelas coletadas nas diferentes velocidades, marcas de bombas, mecanismos de infusão entre as bombas da mesma marca e entre os diferentes tempos de infusão. A principal variação encontrada foi relacionada com o potássio livre, devido à variação inerente aos próprios concentrados, o que foi evidenciado ao se compararem os valores obtidos nas amostras controle e nas amostras colhidas nos tempos médio e final em cada bomba. De acordo com os resultados obtidos, não houve alteração estatisticamente significativa do percentual de hemólise após a passagem pelas bombas de infusão nos modelos analisados.

The lyses of red blood cells, both for immune or non-immune reasons, can cause an elevation in plasmatic hemoglobin, inducing harmful effects mainly in the kidneys and cardiovascular system. This work aims at identifying the possible hemolytic effect, related to mechanical trauma due to the use of different models of infusion pumps, on red blood cells transfused after up to ten days of storage. Three models of pumps were studied (Nutrimat, Infusomat Compact and Volumed), (four devices of each model) with three infusion speeds being tested (120 mL/h, 240 mL/h and 360 mL/h). The parameters used to report the degree of hemolysis were the percentile of hemolysis, hemoglobin levels and free potassium in the plasma. Samples were collected before infusion of red blood cells using the pumps, half way through infusion (1/2 T) and at the end of infusion (T), for the three different speeds. Significant variations were not seen in the analyzed parameters between the control samples and those collected at different speeds, using different models of pumps and infusion mechanisms and among pumps of the same brand and at different infusion times. The greatest variation found involved free potassium, probably due to the different levels found in red blood cell concentrates seen when control samples were compared to values obtained half way through and at the end of infusion for each pump. In agreement with the obtained results changes in the percentile of hemolysis were not found after infusion of the cells using the different infusion pumps.

Síndromes mielodisplásticas: diagnóstico de exclusão / Myelodysplastic syndromes: diagnosis by exclusion

As síndromes mielodisplásticas são comuns nos indivíduos com idade superior a 60 anos e se apresentam laboratorialmente com macrocitose isolada, anemia, citopenias isoladas ou combinadas e alterações morfológicas na medula óssea. O diagnóstico depende da exclusão de causas não clonais e reversíveis. Especialmente nas fases mais precoces da doença, na ausência de excesso de blastos, sideroblastos em anel ou alteração citogenética clonal, o diagnóstico requer um protocolo de exclusão. A exposição recente a agentes tóxicos ou drogas citostáticas, a deficiência de vitamina B12 e ácido fólico e o uso recente de fatores de crescimento são considerados fatores de exclusão absolutos. O etilismo, a anemia da doença crônica, distúrbios metabólicos, hormonais, auto-imunes e infecções virais devem ser excluídos ou interpretados com cautela. Outras doenças da célula-tronco hematopoética devem ser consideradas, sobretudo na SMD hipocelular. Em alguns casos, um período mínimo de seis meses de seguimento é necessário.

Myelodysplastic syndromes are common in elderly people. Laboratory presentation includes isolated macrocytosis, anemia, isolated or combined cytopenias and dysplastic bone marrow. Diagnosis depends on exclusion of non-clonal and reversible disorders. Especially in lowest grade of the disease, with no blast excess, no ringed sideroblasts, no clonal cytogenetic abnormalities the diagnosis requires an exclusion protocol. Recent exposure to toxin, cytotoxic drugs or growth factor therapy and vitamin B12 or folate deficiency are considered absolute exclusion factors precluding the definite diagnosis. Alcohol abuse, chronic inflammatory states, auto-immune disorders, metabolic dysfunctions, hormonal disorders and viral infections must all be ruled out or interpreted with caution. Some diseases of the pluripotential stem cell must also be considered especially in hypocellular MDS. Moreover, in some cases a 6-month follow-up is necessary before a diagnosis be established.