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 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.

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&lt;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 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.

Increased Expression of Circulating Mir-22 in the Peripheral Blood of Patients with Myelodysplastic Syndromes Was Associated with Poorer Prognosis

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2854-2854
Author(s):  
Yuesheng Meng ◽  
Fanli Hua ◽  
Ying Li ◽  
Song Gao
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Myelodysplastic Syndromes ◽  
Peripheral Blood ◽  
Risk Group ◽  
High Risk Group ◽  
Low Risk ◽  
Expression Levels ◽  
Healthy Donors ◽  
Chi Square Analysis

Abstract The molecular pathogenesis of myelodysplastic syndromes (MDS) has not been completely elucidated. Deregulation of expression of some microRNA has been implicated in hematological disorders including MDS. An oncogenic role for miR-22 was recently suggested in MDS in which the expression of miR-22 was increased. However, there were corroborations showing that it could be a tumor suppressor in acute myeloid leukemia. In this study, we examined the expression levels of circulating miR-22 in the plasma of patients with MDS and evaluated its significance in clinical context. The diagnosis of MDS was made according to the WHO classifications and subtypes included 8 cases of RA, 14 cases of RCMD and 6 cases of RARS/RCMD-RS, 10 cases RAEB1 and 7 cases RAEB2. Twenty cases of healthy donors were enrolled as normal control. Written informed consent for sample collection was obtained from all subjects enrolled. EDTA-anticoagulated peripheral blood samples were collected and centrifuged. Circulating microRNAs were purified from the plasma with miRNeasy serum kit (Qiagen). Expression of miR-22 was reverse- transcripted and measured with stem-loop real time quantitative polymerase chain reaction assay. First of all, we proved that the expression of miR-22 was detectable in the plasma of patients with MDS as well as of healthy donors. Our result showed that the expression levels of circulating miR-22 in MDS patients were higher than that in healthy donors (medians 1.360 vs 1.000, P<0.05), among which 13 patients (37.1%) showed marked increase of miR-22 expression (>2 times). Due to the high heterogeneity of MDS and relatively smaller sample size, the subtypes were grouped into two major categories based on IPSS: the low-risk group (including RA, RCMD and RARS/RCMD-RS, totally 18 cases) and the high-risk group (RAEB1 and RAEB2, 17 cases in total). Relative levels of circulating miR-22 were higher in the high-risk group than in the low-risk group (medians= 1.081 vs 2.317, P<0.05). A follow-up study revealed an association of the expression levels of miR-22 with prognosis. Patients with increased expression of miR-22 had poorer clinical response (Chi-square analysis, P<0.05) and lower overall survival rate (Kaplan-Meier analysis, P<0.05). We compared the levels of miR-22 in the plasma with that of mononuclear cells of bone marrow in some MDS patients and found that the relative levels of miR-22 in the plasma and bone marrow cells were not directly co-related (P>0.05), indicating it could be an independent prognostic factor. In summary, this study confirmed an oncogenic presence of miR-22 in MDS. Given the fact that there is a lack of blastic cells in the bone marrow of patients with low-risk MDS but their plasma or serum is easy to obtain, the detection of circulating miR-22 may be of value as a molecular marker in the prognosis of MDS. 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.

Expression of dlk1 in the Bone Marrow Cells of Patients with Myelodysplastic Syndrome and Its Clinical Significance

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 5042-5042
Author(s):  
Zonghong Shao ◽  
Lanzhu Yue ◽  
Rong Fu ◽  
Lijuan Li ◽  
Erbao Ruan ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Early Diagnosis ◽  
Myelodysplastic Syndrome ◽  
Bone Marrow Cells ◽  
Conflicts Of Interest ◽  
Rt Pcr ◽  
Normal Controls ◽  
Lower Expression ◽  
Bone Marrow Blasts ◽  
Marrow Cells

Abstract Abstract 5042 Objective To investigate the expression of dlk1 gene (delta-like 1) in the bone marrow cells of patients with Myelodysplastic syndrome (MDS), and explore the molecular marker for early diagnosis of MDS. Methods The expression of dlk1 mRNA in the bone marrow cells of cases with MDS, AML and normal controls were measured by RT-PCR, aiming to search for the cytogenetic marker of MDS malignant clone. Results The expression of dlk1 mRNA in bone marrow cells of MDS patients (0.7342±0.3652) was significantly higher than that of normal controls (0.4801±0.1759) (P<0.05), and was significantly positively correlated with the proportion of bone marrow blasts(r=0.467,P<0.05). The expression of dlk1 mRNA significantly increased as the subtype of MDS advanced (P<0.05). Patients with abnormal karyotypes displayed significantly higher expression of dlk1 mRNA (0.9007±0.4334) than those with normal karyotypes (0.6411±0.2630) (P<0.05). Patients with higher expression of dlk1(≥0.8) presented significantly higher malignant clone burden (0.4134±0.3999) than those with lower expression (<0.8) of dlk1 (0.1517±0.3109) (P<0.05). Conclusion dlk1 gene was highly expressed in MDS patients, which increased as the subtype of MDS advanced. The expression of dlk1 mRNA was significantly positively correlated with the proportion of bone marrow blasts. High expression of dlk1 gene suggests high malignant clone burden of MDS. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Immunophenotype Rearrangement in Response to Tumor Excision May Be Related to the Risk of Biochemical Recurrence in Prostate Cancer Patients

2021 ◽  
Vol 10 (16) ◽  
pp. 3709
Author(s):  
Paulius Bosas ◽  
Gintaras Zaleskis ◽  
Daiva Dabkevičiene ◽  
Neringa Dobrovolskiene ◽  
Agata Mlynska ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
High Risk ◽  
Radical Prostatectomy ◽  
Cancer Patients ◽  
Peripheral Blood ◽  
Biochemical Recurrence ◽  
Risk Group ◽  
Laparoscopic Approach ◽  
Low Risk ◽  
Tumor Excision

Background: Prostate cancer (PCa) is known to exhibit a wide spectrum of aggressiveness and relatively high immunogenicity. The aim of this study was to examine the effect of tumor excision on immunophenotype rearrangements in peripheral blood and to elucidate if it is associated with biochemical recurrence (BCR) in high risk (HR) and low risk (LR) patients. Methods: Radical prostatectomy (RP) was performed on 108 PCa stage pT2–pT3 patients. Preoperative vs. postoperative (one and three months) immunophenotype profile (T- and B-cell subsets, MDSC, NK, and T reg populations) was compared in peripheral blood of LR and HR groups. Results: The BCR-free survival difference was significant between the HR and LR groups. Postoperative PSA decay rate, defined as ePSA, was significantly slower in the HR group and predicted BCR at cut-off level ePSA = −2.0% d−1 (AUC = 0.85 (95% CI, 0.78–0.90). Three months following tumor excision, the LR group exhibited a recovery of natural killer CD3 − CD16+ CD56+ cells, from 232 cells/µL to 317 cells/µL (p < 0.05), which was not detectable in the HR group. Prostatectomy also resulted in an increased CD8+ population in the LR group, mostly due to CD8+ CD69+ compartment (from 186 cells/µL before surgery to 196 cells/µL three months after, p < 001). The CD8+ CD69+ subset increase without total T cell increase was present in the HR group (p < 0.001). Tumor excision resulted in a myeloid-derived suppressor cell (MDSC) number increase from 12.4 cells/µL to 16.2 cells/µL in the HR group, and no change was detectable in LR patients (p = 0.12). An immune signature of postoperative recovery was more likely to occur in patients undergoing laparoscopic radical prostatectomy (LRP). Open RP (ORP) was associated with increased MDSC numbers (p = 0.002), whereas LRP was characterized by an immunity sparing profile, with no change in MDSC subset (p = 0.16). Conclusion: Tumor excision in prostate cancer patients results in two distinct patterns of immunophenotype rearrangement. The low-risk group is highly responsive, revealing postoperative restoration of T cells, NK cells, and CD8+ CD69+ numbers and the absence of suppressor MDSC increase. The high-risk group presented a limited response, accompanied by a suppressor MDSC increase and CD8+ CD69+ increase. The laparoscopic approach, unlike ORP, did not result in an MDSC increase in the postoperative period.

Analysis of Early Disease Response in Childhood ALL: Can Peripheral Blood Replace Bone Marrow Analysis?.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2523-2523
Author(s):  
David O'Connor ◽  
Jennifer Jesson ◽  
Melanie Bahey ◽  
Lisa Eyre ◽  
Sarah Lawson
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Peripheral Blood ◽  
Risk Group ◽  
Early Time ◽  
High Risk Group ◽  
Time Points ◽  
Post Induction ◽  
Flow Cytometric ◽  
The Uk

Abstract Abstract 2523 The detection of minimal residual disease (MRD) is highly predictive of outcome in pediatric acute lymphoblastic leukemia (ALL) and has now been integrated into the majority of treatment regimens. Early studies demonstrated the strong prognostic value of post-induction MRD measurement, using both molecular and flow cytometric techniques, allowing treatment intensity to be tailored to response. More recent studies have examined the value of MRD measurement at earlier time points and using peripheral blood (PB) in place of bone marrow (BM) in an attempt to reduce invasive procedures. Within the United Kingdom (UK), previous national trials have shown that post-induction MRD assessment, using molecular techniques, is highly predictive of relapse, allowing treatment stratification based on MRD levels. We have previously demonstrated the value of flow cytometric MRD assessment at early time points in response prediction (Motwani et al, Blood 2007a), however, as yet there has been no assessment of MRD in peripheral blood at early time points using flow cytometric methods in the context of a UK national trial. To address this we assessed MRD by flow cytometry using Day 8 (D8) and/or D15 paired PB and BM samples taken from children being treated for ALL at our unit on the UK MRC UKALL2003 trial or ALLR3 relapse trial. Samples were analyzed by 4-color flow cytometry using a large panel of monoclonal antibodies to identify all leukemia-associated immunophenotypes (LAIP). For follow up samples a minimum of 2 markers were identified for each patient and sequential gating was used for analysis. 500 000 to 106cells were acquired for each antibody combination for analysis to be sufficiently sensitive (0.01%). The number of residual leukemic cells was calculated as the percentage of blasts present within total nucleated cells counted. Sensitivity experiments were performed using dilutions of leukemic blasts in normal bone marrow or peripheral blood and demonstrated a sensitivity of 0.01%. Fifty paired BM and PB samples from 46 patients (41 newly diagnosed, 5 relapse) were included in the study (Age 2–22 years, mean 7.8 years). Of these, 22 were taken at D8 and 28 at D15. Forty-two samples were from patients with B-cell ALL, whilst 8 were from patients with T-cell ALL. The most frequent LAIPs used to identify blasts in B-ALL were CD58 (90% of cases), CD45 (75%), CD38 (86%), CD123 (77%). In T-ALL the most common LAIP combinations were CD34 (75%), CD5/CD56 (62.5%), CD5/CD2 (50%) and CD5/CD99 (50%). In keeping with previous reports, B-ALL MRD levels in PB blood were approximately 1.3-log lower than BM MRD levels (D8 mean 2.27% vs. 27.8%; D15 levels 0.63% vs. 7.99%). T-ALL levels were approximately 0.5-log lower. Importantly, the level of MRD in BM and PB in B-ALL showed a strong correlation (r=0.77). Whilst there were insufficient relapses to allow correlation with long-term outcome, PB MRD levels at D8 showed good prediction of post-induction molecular MRD, the current time point used for risk stratification in the UK trial. In patients with PB MRD >1% at D8, 100% (5/5) were in the high-risk group post-induction, significantly more than those with PB MRD <1% at D8, of which only 36.4% (4/11) were allocated to the high risk group post-induction (p=0.023). MRD levels in BM at D8 did not show similar predictive value. Importantly, this early prediction of risk could offer an early opportunity to intensify treatment in this high-risk group, potentially enhancing disease clearance during the induction phase. This study further supports that early assessment of MRD using flow cytometric techniques enhances treatment stratification in pediatric ALL. Furthermore, it shows that PB sampling may provide a markedly less invasive means of MRD assessment in these patients. Validation in a larger cohort of patients is warrented. Disclosures: No relevant conflicts of interest to declare.

Hedgehog Ligands and Receptors Are Deregulated in Myelodysplastic Syndromes

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3817-3817
Author(s):  
Juliana M. Xavier ◽  
Amanda I Dias ◽  
Paulo Latuf-Filho ◽  
Fabiola Traina ◽  
José Vassallo ◽  
...  
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Myelodysplastic Syndromes ◽  
Target Genes ◽  
Conflicts Of Interest ◽  
Megaloblastic Anemia ◽  
Low Risk ◽  
Hedgehog Pathway ◽  
Hematopoietic Stem ◽  
Healthy Donors

Abstract Abstract 3817 Background: The Hedgehog pathway has an important role in self-renew of normal and leukemic stem cells and is upregulated in myeloid leukemias, however there are no studies regarding the Hedgehog pathway in myelodysplastic syndromes (MDS). Hedgehog ligands (Sonic hedgehog [SHh], Indian hedgehog [IHh], and Desert hedgehog [DHh]) are produced by stromal cells and bind to the receptor Patched (PTCH). This binding causes activation of Smoothened (SMO) receptor, resulting in downstream transcription of target genes. Aim: To evaluate Hedgehod pathway components in MDS. Methods: Bone marrow (BM) samples were collected from 39 MDS (25 low risk, 14 high risk-WHO 2008) patients and 26 healthy donors (HD). Relative expressions of PTCH and SMO were obtained by Real Time PCR. For immunohistochemistry, BM biopsies were collected from 21 MDS (17 low risk and 4 high risk) patients and 7 megaloblastic anemia (MA) were used as control. The BM sections were stained with antibodies for DHh, SHh and c-Kit and the percentage of stained nucleated cells was based on an average of 4 high-powered fields. Results: Hedeghog receptors PTCH and SMO were overexpressed in MDS BM cells compared to normal BM as follows; PTCH: [median (min-max)] healthy donors= 2.23 (0.42–9.22); low risk= 6.09 (0.41–25.28); high risk= 3.97 (0.42– 21.01); HD vs low risk and HD vs high risk p= 0.02; SMO: healthy donors= 2.33 (0.00–16.11); low risk= 14.79 (1.89– 41.93); high risk= 34.44 (8.04– 164.28)]; HD vs low risk and HD vs high risk p<0.001. Immunohistochemistry assays showed a higher expression of Hedgehog ligands in MDS bone marrow compared to control. For DHh, the number of stained cells in low risk MDS were higher than in MA [MA = 102(84 –183), low risk= 196(69 – 317) high risk 159(131 – 236)]; MA vs low risk p=0.018 and MA vs high risk p= 0.11. Furthermore, SHh staining was 2 and 2.5 fold higher in low and high risk MDS, respectively, compared to MA [MA= 47(18 – 74), low risk 98.25(53 – 129), high risk 116.25 (93 – 125)] MA vs low risk p= 0.001, MA vs high risk p=0.01. Similar to SHh, c-Kit staining was higher in MDS cases as follows: MA= 95.3(61 – 114), low risk= 106(57–136), high risk= 95.3(61 – 114). Interestingly, a high correlation was found between c-kit and DHh (p=0.002, r=0.63) or c-kit and SHh (p=0.001, r=0.7). Conclusion: To our knowledge, this is the first study of the Hedgehog pathway in MDS. We observed overexpression of Hedgehog receptors and ligands. Furthermore, there is a correlation between these ligands and a marker for hematopoietic stem/progenitors cells. According to these data, we propose that deregulation of Hedgehog in MDS may occur in abnormal progenitors found in MDS bone marrow. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Comparative Value of the Assessment of Minimal Residual Disease in Peripheral Blood at Days 8 and 15 By Flow Cytometry in Childhood Acute Lymphoblastic Leukemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5270-5270
Author(s):  
Marie Loosveld ◽  
Vanessa Nivaggioni ◽  
Isabelle Arnoux ◽  
Denis Bernot ◽  
Chantal Fossat ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
High Risk ◽  
Minimal Residual Disease ◽  
Peripheral Blood ◽  
Residual Disease ◽  
Risk Group ◽  
Lymphoblastic Leukemia ◽  
Low Risk ◽  
The Impact ◽  
Minimal Residual

Abstract Introduction Acute lymphoblastic leukemia (ALL) is the most frequent cancer in childhood, but treatments' progress now allowsto obtain prolonged remission or curein over 90% of the patients. Consequently, therapeutic de-escalation is now an objective for future treatment protocols, providing that biomarkers allow to reliablyidentifygood responders. Among such indicators, low levels of Minimal Residual Disease (MRD) obtained early after induction chemotherapy stand out as good candidates. The latter can be investigated usingmultiparameterflow cytometry (MFC) or real-time polymerase chain reaction (RT-PCR) for immunoglobulins or T-cell receptors (IG TCR) rearrangements. In this study we report the impact on survival of two early points of peripheral blood (PB) MRD assessment by MFC at days 8 and 15 on a cohort of 125 children with B-ALL enrolled in the French FRALLE trial and compared to molecular MRD in the bone marrow (BM) at day 35. Patients and methods. The study enrolled 67 boys and 58 girls and the duration of the study allowed for a median follow up of 52,1months. Median age at diagnosis was 57 months old (range 18 to 196), 101 children were between 1 to 10 years old and 24 were older than 10. Complete blood counts (CBC) at diagnosis showed a median of 6.7x109/L leucocytes (range 0.47 - 151x109/L) and 33% blasts (range 0 to 97%). One hundred and eight children had less than 50x109/L leucocytes while 17 had higher counts. EGIL classification at diagnosis allowed to classify patients as three B-I, 94 B-II, 27 B-III and 1 B-IV. Cytogenetic analyses were performed for 118 patients who were partitioned as follows: low risk n=47, intermediate risk n=55 and high risk n=16 (Harrisson CJ et al., BJH, 2010). Eighty-three patients were in the low risk group and 42 in the high-risk group as described by the FRALLE protocol. Seven patients of the 64 tested had an IKZF1 deletion. During the duration of the study, 20 patients relapsed and 8 died. Corticosensitivitywas defined by less than 1x109/L PB blasts on day 8 andchemosensitivity by less than 5% BM blasts on day 21 on BM smears. PB MRD was assessed in MFC with a single five or ten colors tube adapted to each patient's leukemia associatedimmunophenotypeon a backbone of CD45, CD19, CD10 and CD38. Statistical analyzes examined factors impacting disease-free survival (DFS) using Log rank test and Kaplan-Meier using theMedcalc® software (Ostend, Belgium). P values <0.05 were considered significant. Results None of diagnosis features had any significant impact on DFS: age (p=0,95), risk group (p=0,17), EGIL classification (p=0,55), cytogenetics (p= 0,87), leucocyte count (p=0,36) nor IKZF1 deletion (p=0,2). Of the 125 patients, 9 were corticoresistant, 79 corticosensitive and 37 not evaluable because of less than 1x109/L leucocyte at diagnosis.Corticosensitivity had no impact on DFS (p=0,11). Conversely,chemosensitivity had a significant positive impact on DFS (p= 0,009). Day 8 PB MRD did not oultlineany significantly different DFS, whether considering detectable vs undetectable MRD (p=0.65) or MRD levels (logwisefrom >10-1 to <10-4, p=0,22). Conversely, PB MFC at day 15 appeared highly discriminant. Considering notdetectablevs detectable MRD, 4 years DFS was 91,6+3% vs. 67,6+9% p=0,0013 (Figure 1). Further refining the thresholds of MRD logwisedid not modify the significance (p=0.004; Figure 2). Indeed, DFS at 48 months was 61+15 % (n=16) for MRD >10-3, 74+11% ( n=18) for MRD <10-3->10-4 and 92+3% ( n=91) for MRD<10-4. Comparison of PB MFC MRD on day 15 with day 35 BM molecular MRD showed concordance in 72% of the cases (83 negative/negative and 7 positive/positive, 48 months DFS 94.6+2.7% and 38+20% respectively). Eight patients were negative in PB but positive in BM (DFS 62.5+17%).Twenty seven where positive in PB but negative in BM (DFS 83.5+7.6%).These differences were statistically highly significant (p <0.0001). Conclusion This study demonstrates that even in the good prognosis context of childhood ALL, early MRD retains a highly significant prognostic value. It is of importance that this result was obtained not only on day 35 BM but interestingly, even earlier on day 15 PB. This less invasive procedure can easily be applied, especially for children. It should allow to detectgood responders, with MFC MRD levels below 10-4 for whom a de-escalation of chemotherapy could be considered. Conversely, the detection of blasts by MFC in day 15 PB is worrisome. Disclosures No relevant conflicts of interest to declare.

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