Reduced expression of FLIPSHORT in bone marrow of low risk myelodysplastic syndrome

2007 ◽  
Vol 31 (6) ◽  
pp. 853-857 ◽  
Author(s):  
Paula de Melo Campos ◽  
Fabíola Traina ◽  
Adriana da Silva Santos Duarte ◽  
Irene Lorand-Metze ◽  
Fernando F. Costa ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Myelodysplastic Syndrome ◽  
Low Risk

scholarly journals Mutational Characteristics and Changing Pattern from Idiopathic Cytopenia of Undetermined Significance to High-Risk Myelodysplastic Syndrome Stratified By IPSS-R

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3009-3009
Author(s):  
Eun-Ji Choi ◽  
Young-Uk Cho ◽  
Seongsoo Jang ◽  
Chan-jeoung Park ◽  
Han-Seung Park ◽  
...  
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Myelodysplastic Syndrome ◽  
Rna Splicing ◽  
Conflicts Of Interest ◽  
Low Risk ◽  
International Prognostic Scoring System ◽  
Intermediate Risk ◽  
Sequencing Data ◽  
Undetermined Significance

Background: Unexplained cytopenia comprises a spectrum of hematological diseases from idiopathic cytopenia of undetermined significance (ICUS) to myelodysplastic syndrome (MDS). Revised International Prognostic Scoring System (IPSS-R) is the standard tool to assess risk in MDS. Here, we investigated the occurrence, characteristics, and changing pattern of mutations in patients with ICUS and MDS stratified by IPSS-R score. Methods: A total of 211 patients were enrolled: 73 with ICUS and 138 with MDS. We analyzed the sequencing data of a targeted gene panel assay covering 141 genes using the MiSeqDx platform (Illumina). The lower limit of variant allele frequency (VAF) was set to 2.0% of mutant allele reads. Bone marrow components were assessed for the revised diagnosis according to the 2016 WHO classification. Lower-risk (LR) MDS was defined as those cases with very low- or low-risk MDS according to the IPSS-R. Higher-risk (HR) MDS was defined as those cases with high- or very high-risk MDS according to the IPSS-R. Results: Patients with ICUS were classified as very low-risk (39.7%), low-risk (54.8%), and intermediate-risk (5.5%) according to the IPSS-R. Patients with MDS were classified as LR (35.5%), intermediate-risk (30.4%), and HR (34.1%). In the ICUS, 28 (38.4%) patients carried at least one mutation in the recurrently mutated genes in MDS (MDS mutation). The most commonly mutated genes were DNMT3A (11.0%), followed by TET2 (9.6%), BCOR (4.1%), and U2AF1, SRSF2, IDH1 and ETV6 (2.7% for each). IPSS-R classification was not associated with mutational VAF and the number of mutations in ICUS. In the 49 LR MDS, 28 (57.1%) patients carried at least one MDS mutation. The most commonly mutated genes were SF3B1 (20.4%), followed by TET2 (12.2%), U2AF1 (10.2%), DNMT3A (10.2%), ASXL1 (10.2%), and BCOR (6.1%). Higher VAF and number of mutations were observed in LR MDS compared to ICUS patients. In the 42 intermediate-risk MDS, 27 (64.3%) patients carried at least one MDS mutation. The most commonly mutated genes were ASXL1 (23.8%), followed by TET2 (21.4%), RUNX1 (16.7%), U2AF1 (14.3%), DNMT3A (14.3%), SF3B1 (9.5%), and SRSF2, BCOR, STAG2 and CBL (7.1% for each). In the 47 HR MDS, 36 (76.6%) patients carried at least one MDS mutation. The most commonly mutated genes were TET2 (25.5%), followed by DNMT3A (14.9%), TP53 (14.9%), RUNX1 (12.8%), U2AF1 (10.6%), ASXL1 (10.6%), and SRSF2 and KRAS (6.4% for each). As the disease progressed, VAF and number of the MDS mutations gradually increased, and mutations involving RNA splicing, histone modification, transcription factor or p53 pathway had a trend for increasing frequency. Specifically, ASXL1, TP53, and RUNX1 mutations were the most striking features in patients with advanced stage of the disease. Cohesin mutations were not detected in ICUS, whereas these mutations were detected at a relatively high frequency in HR MDS. Our data were summarized in Table 1. Conclusions: We demonstrate that on disease progression, MDS mutations are increased in number as well as are expanded in size. Furthermore, a subset of mutations tends to be enriched for intermediate- to HR MDS. The results of this study can aid both diagnostic and prognostic stratification in patients with unexpected cytopenia. In particular, characterization of MDS mutations can be useful in refining bone marrow diagnosis in challenging situations such as distinguishing LR MDS from ICUS. Disclosures No relevant conflicts of interest to declare.

FLIP (Long) and FLIP (Short) Expression in Bone Marrow Cells in Patients with Myelodysplastic Syndrome: Correlation with FAB and WHO Classification.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4926-4926
Author(s):  
Paula Campos ◽  
Fabiola Traina ◽  
Adriana Duarte ◽  
Bruno Benites ◽  
Marcelo Brandao ◽  
...  
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Myelodysplastic Syndrome ◽  
Mrna Expression ◽  
Who Classification ◽  
Fas Ligand ◽  
Low Risk ◽  
World Health ◽  
Caspase 8 ◽  
Marrow Cells

Abstract The paradox of peripheral cytopenias despite of normo/hypercellular marrow in myelodysplastic syndrome (MDS) has been ascribed to excessive intramedullary hematopoietic cell apoptosis. Several apoptosis-inducing systems, including Fas/Fas ligand and TNF-related apoptosis-inducing ligand (TRAIL) and its receptors, are upregulated in MDS. FLIP (FLICE (FAS-associated death-domain-like IL-1β-converting enzyme)-inhibitory protein) was identified as a FAS and TRAIL signal inhibitor. The largest variant FLIPLong (FLIPL) was originally characterized as a molecule with inhibitory activity on caspase-8. The short splice form termed FLIPShort (FLIPS) has also been characterized as a potent (TRAIL-induced) apoptosis inhibitor. However, whereas FLIPL and FLIPS have been described as death receptor pathway inhibitors, recent data suggest that physiologically, FLIPL may have caspase-8-activating properties. This study aims to characterize the expression of FLIPL and FLIPS based on mRNA, by Real-time quantitative PCR, in marrow cells from MDS patients and to correlate the expression with French-American-British (FAB) and World Health Organization (WHO) classification. For each sample, results were first calculated as a ratio of the total transcript number of FLIPL or FLIPS and the total transcript number of the endogenous reference gene (β-actin) to obtain a normalized target value. Transcript ratios of each sample were normalized against the respective ratio of a pool of 6 normal bone marrow donors (NBM), and the ratio between the two was used as measure for the relative FLIPL or FLIPS level. We hypothesized that FLIPL and FLIPS expression differed between low and high risk of MDS. Marrow aspirates were obtained from 6 NBM and 16 patients with MDS out of treatment (7 males, 9 females; 23–78 (median 64) yo). The National Ethical Committee Board approved this study, informed-written consent was obtained from all patients and donors. According to FAB classification, patients were distributed as: 10 RA, 2 RARS and 4 RAEB. According to WHO classification: 10 RCMD, 2 RCMD-RS, 3 RAEB-1 and 1 RAEB-2. FLIPS mRNA expression were significantly higher in high risk DS according to FAB and WHO classification; RA/RARS compared with AREB (0.08 [0.0–2.3] vs 0.67 [0.36–1.54]; P = 0.03); RCMD and RCMD-RS compared with RAEB-1 and RAEB-2 (0.08 [0.0–2.3] vs 0.67 [0.36–1.54]; P = 0.03). However, FLIPL mRNA expression also tended to be higher in high risk MDS according to FAB and WHO classification, though not significantly different: RA/RARS compared with AREB (1.18 [0.06–3.43] vs 1.65 [0.51–3.63]; P = 0.46); RCMD and RCMD-RS compared with RAEB-1 and RAEB-2 (1.18 [0.06–3.43] vs 1.65 [0.51–3.63]; P = 0.46). Lower FLIPS level in low risk MDS marrows, in addition to the well described upregulation of extracellular proapoptotic signals, would explain the increased susceptibility of hematopoietic cells in low risk MDS marrow to death-inducing stimuli. The fact that FLIPL expression did not differ according to FAB and WHO classification could be related to the hypothesis that FLIPL may have caspase-8-activating properties rather than anti-apoptotic activity. Differential regulation of FLIPL and FLIPS according to risk groups in MDS patients might result in different rates of apoptosis. Further studies are needed to elucidate the mechanisms controlling and regulating FLIP expression in normal and malignant hemopoietic cells.

Flow Cytometric Detection of Human Telomerase Reverse Transcriptase Expression in CD34 Positive Precursor Fraction in Myelodysplastic Syndrome Bone Marrow.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4300-4300
Author(s):  
Hiroshi Handa ◽  
Takafumi Matsushima ◽  
Norifumi Tsukamoto ◽  
Masamitsu Karasawa ◽  
Hiroyuki Irisawa ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
High Risk ◽  
Myelodysplastic Syndrome ◽  
Disease Progression ◽  
Risk Group ◽  
Telomerase Activity ◽  
Low Risk ◽  
Htert Expression ◽  
Trap Assay

Abstract Telomerase activity has been found in most common cancers indicating that telomerase detection may be a useful marker in cancer diagnosis. For detection of telomerase activity and the expression of associated genes in cells, TRAP assay and RT-PCR are customarily used. Immunohistochemical detection of hTERT is useful to detect telomerase-positive cells in a background of non- cancerous cells. We developed a method for the detection of intra-nuclear hTERT protein, in a sub-population of hematopoietic cells, using concurrent staining cell surface antigen and multi color flow cytometry. Human leukemia and myeloma cell lines showed 100% positivity, whereas neutrophils of normal subjects showed 0% positivity, it is consistent with telomerase activity assessed by TRAP assay (r=0.71, p<0.0001) and previous observations. Then we applied this method to analyze hTERT expression in myelodysplastic syndrome (MDS). Forty MDS patients samples were obtained, 36 patients were diagnosed as low risk MDS (RA), 14 patients were diagnosed as high risk MDS (RAEB or RAEB-t) according to FAB classification. All samples were acquired after informed consent was obtained from the patients. Expression of hTERT protein was higher in CD34-positive blast-gated cells than CD34-negative blast-gated cells. The percentage of the CD34+ cells expressing hTERT ranged from 9.66% to 90.91% in low risk MDS patients, whereas from 50.46% to 97.68% in high risk MDS. The expression level was higher in the high risk group compared to that in the low risk group in MDS (p=0.0054, p=0.0084). This observation implied that telomerase up-regulation and hTERT expression were important for disease progression and could be a marker of more advanced disease. In subsets of MDS and AML bone marrow specimens obtained from these patients, we examined the hTERT expression in CD34+/CD38 high cells and CD34+/CD38 low cells containing stem cell fraction. Of interest, some of the patients showed higher expression of hTERT in CD34+/CD38 low cells than in CD34+/CD38 high cells. This observation is inconsistent with previous reports describing normal bone marrow hematopoietic cell findings. We speculated that this phenomenon could be a marker of MDS abnormality and that telomerase up-regulation may be initiated in the more primitive precursor fraction containing hematopoietic stem cells during the disease progression. Telomerase studies may be useful for definition of the risks associated with disease severity. Multi-parameter nature of flow cytometry and its ability to identify cellular sub-populations will facilitate a fuller understanding of the mechanisms of activation of telomerase.

scholarly journals GDF11 Increased in Patients with Myelodysplastic Syndrome

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 5224-5224
Author(s):  
Yu Han ◽  
Huaquan Wang ◽  
Zonghong Shao
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Myelodysplastic Syndrome ◽  
Peripheral Blood ◽  
Conflicts Of Interest ◽  
Risk Group ◽  
Low Risk ◽  
Mean Corpuscular Hemoglobin ◽  
Corpuscular Hemoglobin ◽  
Normal Controls

Abstract Objective To analyze the concentration of growth differentiation factor 11(GDF11) in peripheral blood of patients with myelodysplastic syndrome (MDS), so as to evaluate the relationships between these changes and erythropoiesis functions and to explore the role of GDF11 in the pathogenesis of MDS. Methods The concentration of GDF 11 in peripheral blood was detected by enzyme-linked immuno sorbent assay in 44 MDS patients and 10 normal controls from September 2014 to June 2015 at our hospital. The percentage of nucleated erythrocyte (CD235a) in bone marrow was detected by flow cytometry. The correlation between these changes and erythropoiesis functions, including red blood cell count, hemoglobin, reticulocyte (RET%), hematokrit (Hct), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular-hemoglobin concentration (MCHC) and late erythroblast in bone marrow were evaluated. Results (1)The concentration of GDF11(128.67±47.62)in high-risk MDS patients was significantly higher than that of low-risk MDS patients (65.96±36.55,p<0.01)and higher than that of normal controls (29.76±10.10,p<0.01); The concentration of GDF11 in low-risk MDS patients was significantly higher than that of normal controls (p<0.05). (2) The expression of CD235a in high-risk group(38.49±5.42)was not different with that in low-risk group(42.64±7.36, p>0.05). (3)In high-risk MDS patients, the expression of GDF11 was negatively correlated with Hb, RET%, RBC, MCHC, Hct in peripheral blood and late erythroblast, CD235a+ cells in bone marrow(r=-0.437,r=-0.428,r=-0.444,r=-0.553,r=-0.661,r=-0.436,r=-0.52,all p<0.05),and the expression of GDF11 was positively correlated with MCV(r=0.52, p <0.05),but it was not correlated with MCH (p >0.05).(4) In low-risk MDS patients, the expression of GDF11 was negatively correlated with Hb, RET% (r=-0.491Ar=-0.606,both p<0.05),it was not correlated with RBC, MCHC, MCV, MCH, Hct, late erythroblast and CD235a+ cells (all p>0.05). Conclusion GDF11 increased in patients with MDS and it was negatively correlated with late erythropoiesis. Disclosures No relevant conflicts of interest to declare.

scholarly journals Genetic Mutation in Cytopenic Patients: Distinctive Genomic Profile between Preclinical Vs. Clinical Myelodysplastic Syndrome

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5429-5429
Author(s):  
Kritanan Songserm ◽  
Amornchai Suksusut ◽  
Sunisa Kongkiatkamon ◽  
Kitsada Wudhikarn ◽  
Chinnachote Teerapakpinyo ◽  
...  
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Myelodysplastic Syndrome ◽  
Somatic Mutations ◽  
Conflicts Of Interest ◽  
Gene Mutations ◽  
Genetic Alterations ◽  
Genetic Mutation ◽  
Low Risk ◽  
Genomic Profile

Genetic mutation in cytopenic patients: Distinctive genomic profile between preclinical vs. clinical myelodysplastic syndrome. Introduction Myelodysplastic syndromes (MDS) are heterogeneous groups of clonal hematopoietic disorders. The current diagnosis of MDS is based on morphologic assessments of dysplasia which are subjected to inter-observer variability and cytogenetic abnormalities which are frequently absent. Somatic mutations in myeloid-related genes have been identified in MDS. However, they are also found in idiopathic cytopenia of unknown significance (ICUS) that shows no significant dysplasia. Therefore, we aimed to explore the clinical implications of genetic mutations in ICUS and compared with MDS. The secondary objective was to find association between degree of dysplasia and somatic mutations. Materials and Methods The patients with peripheral cytopenia ≥1 lineage (ANC < 1,800/mm3, hemoglobin < 10 gm/dL, platelet < 100x109/mL) without explainable causes were enrolled. Bone marrow aspirates were evaluated independently by 2 hematologists. Of note, dysplasia are defined by WHO 2008 classification (eg. Erythroid lineage: ring sideroblasts, megaloblastoid change; granulocytic lineage: hypogranularity, pseudopelger-huet anomaly; megakaryocytic lineage: hypolobate, micro-megakaryocyte). The significant dysplasia cut off was 10% in single lineage or more. If there was a discrepancy, the third hematologist would help to reach the final consensus. We extracted DNA from bone marrow and performed next generation sequencing (NGS) that targeted 143 myeloid-related genes. Results Forty-eight patients were enrolled in this study. The median age at diagnosis was 70 years (71-96). Results of bone marrow examinations were categorized by morphology into 3 groups; non-significant dysplasia (dysplasia < 10%) 27%, low risk MDS (IPSS-R ≤3.5) 42% and high-risk MDS/sAML (IPSS-R >3.5/Blast≥20% in BM or peripheral blood) 31%. Most of cases (77%) carried normal cytogenetics while other genetic alterations were complex chromosome (6%), -Y (6%), del(5q) (4%), trisomy 8 (2%), del(20q) (2%), i(17q) (2%). Thirty from 48 cases (62%) harbored more than 1 somatic mutation. Twenty-eight gene mutations were identified. Mutations were detected 1.6 mutation per 1 patient in average. Most frequent somatic mutations were ASXL1:10/80 (12%), TET2:9/80 (11%), MFDS11: 6/80 (7%), TP53:6/80 (7%), and RUNX1:5/80 (6.25%). The proportions of cases with somatic mutations were not different across the groups (no dysplasia 50%, non-significant dysplasia 80% and significant dysplasia 62%). According to mutation types in each group, mutations in epigenetic pathways were the most frequent mutations across all patient subgroups (ICUS 64.7%, low-risk MDS 51.8 %, and high-risk MDS 52.5%). Mutations in transcription factor were predominated in MDS (18.5% and 25.0% in low-risk and high-risk MDS, respectively) compared to ICUS (11.7%). Individual average frequency of gene mutations was significantly different between disease subtype (high risk MDS 2.7 gene/person, low risk MDS 1.1 gene/person, ICUS 1.3 gene/person (P=.038). Higher variant allele frequency (VAF) of mutated genes was significantly observed in high risk MDS (38.3%) compared to low risk MDS (30.8%) and preclinical MDS (29.0%) (P=.03). Conclusion In conclusion, molecular profiling was significantly different between preclinical MDS and MDS groups in terms of types of somatic mutations and VAF. This unique contrast could be used to distinguish between preclinical MDS and clinically significant MDS. In contrast, degree of marrow dysplasia was not associated with number of gene mutations in this study. Prediction for clinical consequent of somatic mutations in CCUS requires long term follow up. Disclosures No relevant conflicts of interest to declare.

scholarly journals Bone Marrow Fibrosis at Diagnosis is Associated with TP53 Overexpression and Adverse Prognosis in Low-Risk Myelodysplastic Syndrome

2017 ◽  
Vol 55 ◽  
pp. S143
Author(s):  
R.P.G. Lemes ◽  
F.B. Duarte ◽  
M.C. Barbosa ◽  
J.P. Vasconcelos ◽  
F.D. Rocha ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Myelodysplastic Syndrome ◽  
Low Risk ◽  
Bone Marrow Fibrosis ◽  
Marrow Fibrosis

scholarly journals Bone marrow fibrosis at diagnosis is associated with TP53 overexpression and adverse prognosis in low-risk myelodysplastic syndrome

2017 ◽  
Vol 181 (4) ◽  
pp. 547-549 ◽  
Author(s):  
Fernando B. Duarte ◽  
Maritza C. Barbosa ◽  
Talyta E. Jesus dos Santos ◽  
Romélia P. G. Lemes ◽  
João P. Vasconcelos ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Myelodysplastic Syndrome ◽  
Low Risk ◽  
Bone Marrow Fibrosis ◽  
Marrow Fibrosis

scholarly journals P53 and CD34 Immunohistochemical Expression on Bone Marrow Biopsy As Predictive Marker of Response to Erythropoietin in Low Risk Myelodysplastic Syndrome

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5511-5511
Author(s):  
Gianluigi Reda ◽  
Francesca Boggio ◽  
Ramona Cassin ◽  
Marta Riva ◽  
Marco Barella ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Myelodysplastic Syndrome ◽  
Treatment Response ◽  
Bone Marrow Biopsy ◽  
Univariate Analysis ◽  
Low Risk ◽  
P Value ◽  
P53 Expression ◽  
Loss Of Response

Abstract Introduction: Erythropoietin stimulating agents (ESAs) are first-line therapy for International Prognostic Scoring System (IPSS)-low risk myelodysplastic syndrome (MDS) when symptomatic anaemia (Hb <10 g/dl) is associated with serum erythropoietin (EPO) <500 mU/mL. Treatment response rate, according to international working group (IWG) response criteria (1- 5), ranges from 40 to 60%. Attempts have been made to build prognostic scores (integrating transfusion need, erythropoietin level, R-IPSS and ferritin) able to predict response to ESAs (6-7). No data concerning the predictive value of morphological and immunohistochemical analysis on bone marrow biopsy in low risk MDS patients are emerged till now. Methods: We retrospectively examined 96 IPSS low/int-1 MDS patients treated with ESAs in order to evaluate the morphological and immunohistochemical features of the bone marrow biopsies performed at baseline. All the patients had hemoglobin (Hb) ≤10 g/dL and serum EPO <500 mU/mL and received EPO alfa or β 40 000-80 000 IU/week for at least 12 weeks. Response to ESAs treatment was evaluated according to IWG 2006 criteria. A detailed analysis of the morphological features (including quantitative and qualitative changes in the erythroid, myeloid and megakariopoyetic lineages) was performed, together with an immunohistochemical evaluation of p53 expression and CD34-positive blasts percentage. Results: Sixty-eight percent of the patients were classified as responder while 32% as non-responder. The morphologic profiles of the responder and non-responder did not differ significantly. A significant correlation was found between the response to the therapy and a percentage of CD34-positive blasts > 3% (univariate analysis: p= .0211). Moreover p53 expression in less than 1% of the nucleated cells correlated with the treatment response (univariate analysis: p = .0086). These results were also confirmed on multivariate analysis (p= .002). Fifty-eight percent of patients maintained response to ESAs for the entire duration of the follow up while 42% lost the response (median follow up: 35 months). More than 3% CD34-positive blasts was associated with a higher probability to lose the response to ESAs (p = .00003). Kaplan-Meier method was than applied to estimate the response duration. Patients with more than 3% CD34-positive blasts showed earlier loss of response in comparison with those with a lower percentage of blasts (p value= .0003; HR=3.9, IC 95% 1.8154-8.6238). Finally, the expression of p53 in more than ≥ 1% of the nucleated cells correlated with the loss of response before 24 mounts (p value= 0.029). Conclusions: Our study identifies 2 well-known parameters that could be useful to better predict outcome of ESAs therapy in low risk MDS patients. Therefore role of bone marrow biopsy together with its diagnostic contribution in the clinical evaluation of MDS patients (cellularity, morphologic dysplasia, percentage of CD34-positive blasts, grading of bone marrow fibrosis) could be helpful as a predictive tool in ESAs candidate patients. Further studies based on larger series of patients are needed to confirm these preliminary results. Disclosures Reda: Gilead: Consultancy; ABBVIE: Consultancy; Janssen and Cilag: Consultancy; Celgene: Consultancy. Riva:Jannsen and Cilag: Consultancy; Novartis: Consultancy; Celgene: Consultancy. Molteni:Janssen and Cilag: Consultancy; Novartis: Consultancy; Italfarmaco: Consultancy; Celgene: Consultancy; AMGEN: Consultancy. Cortelezzi:abbvie: Consultancy; novartis: Consultancy; roche: Consultancy; janssen: Consultancy.

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