Bone Marrow Fibrosis in Patients with Inherited Bone Marrow Failure Syndromes.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3192-3192
Author(s):  
Neelam Giri ◽  
Irina Maric ◽  
Robert Wesley ◽  
Diane C. Arthur ◽  
Pierre Noel ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Prognostic Significance ◽  
P Value ◽  
Fisher Exact Test ◽  
Bone Marrow Fibrosis ◽  
Bone Marrow Biopsies ◽  
Bone Marrow Failure Syndromes ◽  
Marrow Fibrosis

Abstract Abstract 3192 Poster Board III-129 Introduction Bone marrow fibrosis has been reported in many benign and malignant disorders, and it may be associated with a poor prognosis in patients with chronic idiopathic myelofibrosis and adult myelodysplastic syndrome (MDS). There are no data on the incidence or significance of bone marrow fibrosis in patients with the inherited bone marrow failure syndromes (IBMFS), genetic disorders characterized by cytopenias, distinctive clinical features, varied molecular pathways and high risks of MDS and acute myeloid leukemia. We have now studied marrow fibrosis in the four most common IBMFS: Fanconi anemia (FA), Diamond-Blackfan anemia (DBA), dyskeratosis congenita (DC), and Shwachman-Diamond syndrome (SDS). Patients and Methods Blinded bone marrow biopsies were analyzed from 42 patients: 12 FA, 13 DBA, 13 DC, and 4 SDS. Reticulin fibrosis was graded on a scale of 1-4 according to the quantity and pattern of distribution of reticulin. The frequencies of abnormalities in marrow fibrosis, cellularity, MDS, cytogenetic clones, blood counts, erythropoietin levels and treatment were compared between the disorders. Fisher exact test was used to compare frequencies and two-sided Wilcoxon rank sum test was used to compare continuous variables. P value of less than 0.05 was considered statistically significant for all tests. Results See Table. Patients with FA, DBA and DC were older than those with SDS; there was an excess of females with FA and males with DC and DBA. Patients with FA, DC and SDS had multilineage cytopenias, while most patients with DBA had only anemia. All patients with FA and DC had bone marrow hypocellularity; it was less frequent in those with DBA or SDS (P<0.001). A significantly higher proportion of patients with FA or DC (75%) had increased reticulin fibrosis (grade 2 or 3) compared with DBA or SDS (P=0.002). Most patients had grade 2 fibrosis; only 2 patients with FA had grade 3 fibrosis, and none had grade 4 fibrosis. In a multivariate analysis, bone marrow fibrosis correlated significantly with the presence of thrombocytopenia (P=0.01) and neutropenia (P=0.01), but not with anemia (P=0.8). The presence of fibrosis correlated significantly with the presence of high erythropoietin levels (P=0.006) and the presence of hypocellularity (P=0.003); contrary to expectations, even very hypocellular marrow biopsies had increased reticulin. Fibrosis did not correlate with the need for treatment, morphologic MDS (P=0.7) or cytogenetic abnormalities (P=0.2). Conclusion This is the first report of bone marrow fibrosis in patients with an IBMFS. The frequency of grade 2 or 3 bone marrow fibrosis was as high as 75% in FA and DC. There was a direct correlation between the presence of marrow hypoplasia and fibrosis in patients with FA and DC. Longitudinal studies are required to determine the prognostic significance of increased marrow fibrosis in patients with an IBMFS. Disclosures No relevant conflicts of interest to declare.

Alterations In Wnt Signalling In the Megakaryocytic Lineage Leads to Bone Marrow Failure and Myelofibrosis

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 628-628
Author(s):  
Simon D Calaminus ◽  
Gareth Inman ◽  
Cedric Ghaevert ◽  
Owen Sansom ◽  
Steve P Watson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Wnt Pathway ◽  
Bone Marrow Failure ◽  
Prognostic Score ◽  
Fold Increase ◽  
Wnt Signalling ◽  
Beta Catenin ◽  
Bone Marrow Fibrosis ◽  
Platelet Factor ◽  
Marrow Fibrosis

Abstract Abstract 628 Introduction: Wnt signalling is fundamental in controlling stem cell self-renewal, cell proliferation and development in multicellular organisms. Stabilization of beta catenin or loss of the scaffold protein adenomatis polyposis coli (APC) causes aberrant activation of wnt signalling and often leads to cancer. Mutations to wnt pathway members in haematopoietic stem cells leads to haematopoietic failure and rapid lethality. In this study, we demonstrate that aberrant wnt signalling in the megakaryocyte lineage underlies myelofibrosis. Methods: We created a series of mice with altered wnt pathway signalling in their megakaryocytic lineage using PF4-Cre (platelet factor 4 cre) as follows: Ctnnb1fx(ex3)/wt_ Expresses stabilized active beta catenin henceforth termed PF4bcat+; APCfx/fx_ Loss of APC stabilises beta catenin termed PF4APC-KO; Ctnnb1fx/fx b-catenin knockout termed PF4bcat-KO. Results: By day 40, PF4bcat+ and PF4APC-KO mice are severely underweight, anaemic (wt 6.8+0.2×106/ml v. PF4bcat+ 4.2+0.3×106/ml, PF4APC-KO 5.5+0.5×106/ml), and have a significant reduction in platelet number (wt 1033+37×106/ml v. PF4bcat+ 717+57×106/ml, PF4APC-KO 747+68×106/ml). Furthermore PF4bcat+ and PF4APC-KO mice develop bone marrow fibrosis and consistently die within 50 days of birth. Both populations of mice have splenic extramedullary haemopoiesis with hyperplasia of splenic megakaryocytes, leading to a dramatic increase in spleen to body size ratio. In addition, both PF4bcat+ and PF4APC-KO mice have increased peripheral blood levels of active TGFb, providing a likely molecular basis of the induction in bone marrow fibrosis. PF4APC-KO and PF4bcat+ mutant mice show a dramatic (>10-fold) increase in platelet b-catenin protein levels over wt samples. By comparison, human myelofibrosis patients (n=16) show a 2.7+0.6-fold increase in platelet b-catenin expression over controls. Moreover, overexpression of b-catenin within human patient samples is correlated with a worsening Primary Myelofibrosis prognostic score (Blood, 2009 vol 113 p. 2895), with those patients with a low score (n=7) having a 1.3+/−0.37-fold increase over control, intermediate score (n=4) 4.52+/−1.23-fold, and high score (n=1) 4.56-fold. In contrast PF4bcat-KO mice show no changes in whole blood counts, weight, or evidence of splenic extramedullary haematopoiesis, indicating that b-catenin removal does not adversely affect megakaryocyte development or function. Conclusions: Stabilisation of b-catenin within mouse megakaryocytes leads to a myelodysplastic disorder and myelofibrosis. This finding demonstrates a defined role for aberrant activation of the wnt signalling pathway and marks the wnt pathway and the megakaryocyte lineage as important potential drug targets for the treatment of myelodysplastic disorders. Disclosures: No relevant conflicts of interest to declare.

Significance of Bone Marrow Karyotype and Morphology in Patients with Inherited Bone Marrow Failure Syndromes,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3431-3431
Author(s):  
Neelam Giri ◽  
Blanche P Alter ◽  
Helkha Peredo-Pinto ◽  
M. Tarek Elghetany ◽  
Irina Maric ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Disease Progression ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Myelogenous Leukemia ◽  
Monosomy 7 ◽  
Cytogenetic Abnormalities ◽  
Bone Marrow Failure Syndromes ◽  
Number Of Patients

Abstract Abstract 3431 Recurring clonal cytogenetic abnormalities have been described in patients with Fanconi anemia (FA) and Shwachman-Diamond syndrome (SDS). In FA, gains of 3q and monosomy 7 (−7) imply progression to myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML). In SDS, isochromosome 7q and deletion (del) 20q are usually benign. Dyskeratosis congenita (DC) and Diamond-Blackfan anemia (DBA) do not have unique clones. We report here the types and frequencies of cytogenetic clones and their association with morphologic MDS or AML in the major inherited bone marrow failure syndromes (IBMFS), in a prospective/ retrospective study of patients with FA, SDS, DC and DBA enrolled in the NCI IBMFS cohort from 2002–2010. Bone marrow (BM) morphology and cytogenetics (G-banding; selected FISH, CGH, SKY) performed at our institute and all outside cytogenetics reports were centrally reviewed. Cytogenetic abnormalities were defined and karyotypes designated according to ISCN (2009). Two independent blinded hematopathologists reviewed BM morphology. Diagnosis of morphologic MDS was based on a modification of WHO 2008 and required ≥10% dysplasia in 2 cell lineages. Data analysis was both cross-sectional and longitudinal. P values are global comparing all 4 disorders using Fisher's exact test.ParameterAll IBMFSFASDSDCDBAP valueTotal number (N)12835113646–N with clone ever2817 (49%)4 (36%)4 (11%)3 (7%)<0.01N with MDS ever105 (14%)3 (27%)1 (3%)1 (2%)0.01N with clone + MDS75 (14%)2 (18%)00<0.01N with clone alone2112 (34%)2 (18%)4 (11%)3 (7%)<0.01N with MDS alone301 (9%)1 (3%)1 (2%)0.3N with clone at 1st BM179 (26%)4 (36%)3 (8%)1 (2%)<0.01N with clones at follow-up118012<0.01N with follow-up BMs591791716–Median follow-up in years3 (0–19)6 (1–16)2 (1–6)3 (0–19)2 (0–10)– More FA and SDS patients had clones and/or MDS compared with DC or DBA (Table). MDS was always associated with clones in FA but not in the other IBMFS. In FA, bone marrow transplant (BMT) or death occurred with similar frequencies in those with or without clones. Among 17 patients with clones, follow-up cytogenetics were unavailable in 5; of these, 2 with clone alone [one with del 7q and 18p and one with t(3;6)(q?25;p?21)] progressed to AML, while one with clone and MDS died from other causes. Recurring abnormalities in 12 FA patients with clones followed for up to 16 years, included gains of 1q in 4, −7 or del 7q in 3, and deletions of 6p, 13q, 18p and 20q in 2 patients each; only one had gain of 3q. These patients showed fluctuation or disappearance of clones, new appearance of clones, stable clone, or clonal evolution. Progression to MDS occurred with gain of 1q and 6p deletion, gain of 3q, or −7 in 3 patients, respectively; one patient with MDS had clonal persistence. No disease progression was seen in 5 FA patients with clone alone. All 5 SDS patients with clones and/or MDS are alive with no disease progression. The 4 with a clone had stable persistent del 20q as a sole abnormality; 2 had MDS and 2 did not. One had MDS with a normal karyotype. Four DC patients had abnormal clones including 2 with gain of 1q only. One patient with 1q gain died from pulmonary fibrosis. Three others are alive; 2 with stable clones at 7 and 19 years' follow-up, respectively. One additional DC patient has morphologic MDS but no clone. All 3 DBA patients with clones had del 16q, 2 alone and 1 with del 9p; none had MDS. The clones were transient in 2, disappearing within 1–2 years; the third was recently identified. None of these had disease progression. One patient with morphologic MDS alone died from complications of iron overload. This study shows that clonal chromosome abnormalities occur more frequently in FA and SDS than in DC and DBA. Gain of 3q in FA was not as common here as reported by others. This is the first comprehensive study of clones and MDS in DC and DBA. Strengths of this study include the large number of patients, and central review of cytogenetics and morphology. It is unbiased compared with FA literature reports that include many patients referred for BMT. Limitations include a relatively small number of patients with each diagnosis and short follow-up in most. The study demonstrates that clones may fluctuate or disappear, and may not per se portend a bad prognosis. Progression to clinically significant MDS or AML may be related to the severity of cytopenias and not to clone alone, and warrants more extensive long-term studies. Disclosures: No relevant conflicts of interest to declare.

Utility of Telomere in Diagnosis of Bone Marrow Failure Syndromes

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4793-4793
Author(s):  
Hasan Ahmed Abdel-ghaffar ◽  
Hosam Zaghloul ◽  
Ahmed El-Waseef ◽  
Mohamed El-Naggar ◽  
Mohamed Mabed ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Telomere Length ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Quantitative Polymerase Chain Reaction ◽  
Control Group ◽  
Hematopoietic Stem ◽  
Relative Telomere Length ◽  
Bone Marrow Failure Syndromes ◽  
Significant Difference

Abstract Background and aim of the work: Bone marrow failure syndromes (BMFS) includes inherited and acquired conditions. Inherited bone marrow failure includes a number of syndromes; with Fanconi anemia (FA) being the most common one of them. Telomeres are eroded with cell division, but in hematopoietic stem cell, maintenance of their length is mediated by telomerase. Short telomeres can result in instability of cell function where diseases occur. Bone Marrow Failure might be developed due to low telomerase activity or short telomeres. Our study is aiming to evaluate the utility of Real Time Quantitative-Polymerase Chain Reaction (RT-qPCR) in measuring the relative telomere length and its significance in diagnosis and prognosis of patients with BMFS. Materials and methods: The study includes 3 groups: A group of congenital BMF (29 patients), a group of acquired BMF (10 patients) and a third control group (15 cases). The relative telomere length is evaluated for them using RT-qPCR. Results: We have found that there is a significant difference in relative telomere length between congenital group and controls (p=0.001), also a significant difference between acquired group and controls (p= 0.029). However, there is no significant difference between congenital and acquired groups (p= 0.479). There is no significant correlation between the telomere length and the overall survival or prognosis of the patients of BMFS. Conclusion: We conclude that the telomere length is significantly altered in patients with BMFS whether being congenital or acquired compared to the control group. Disclosures No relevant conflicts of interest to declare.

Bone Marrow Biopsy and Aspirate Evaluation in 90 Patients with Essential Thrombocythemia Treated with PEG Interferon alpha-2b. Preliminary Results.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1523-1523 ◽  
Author(s):  
L. Gugliotta ◽  
S. Bulgarelli ◽  
A. Tieghi ◽  
S. Asioli ◽  
G. Gardini ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Platelet Count ◽  
Bone Marrow Biopsy ◽  
Essential Thrombocythemia ◽  
Bone Marrow Fibrosis ◽  
Minor Response ◽  
Bone Marrow Biopsies ◽  
Hematological Response ◽  
Second Year ◽  
Marrow Fibrosis

Abstract Ninety patients with Essential Thrombocythemia (ET) where object of a phase II prospective multicentre study designed to evaluate efficacy, safety and tolerability of a two years treatment with PEG Interferon α-2b (PEG Intron, Schering Plough). The patients, 30 M and 60 F, 18–72 years old (median 45), observed in 16 Hematological Institutions of the Gruppo Italiano Malattie Mieloproliferative Croniche (GIMMC), received the ET diagnosis according to the PVSG criteria. At PEG Intron treatment start the patients showed: previous cytoreduction 97% (IFN α 31%), platelet count >1000 x 109/L 81%, splenomegaly 22%. At the end of the first year The PEG Intron starting dose of 25 μg/week resulted increased to a mean value of 55 μg/week and the Hematological Response (HR = Plts <500x109/L) was registered in 79% of the patients still on treatment. At the end of second year 65 patient still receiving PEG Intron (mean dose 31 μg/week) showed a maintenance of the HR (66%), a Partial Response (17%) and a Minor Response (17%). By utilizing the data included in the study CRFs we preliminarily evaluated the bone marrow biopsy and aspirate both performed at baseline, after 1 and 2 years in 89 and 86, 79 and 67, 57 and 50 patients, respectively. Data concerning the bone marrow biopsies after 1 year of treatment are reported: BONE MARROW BIOPSY BASELINE % 1 YR % 2 YRS % Cellularity increased 56 51 48 Granulopoiesis increased 51 54 39 Erytropoiesis increased 29 24 23 MK number increased 99 90 84 MK size increased 78 69 62 MK ploidy 54 51 42 MK dystrophy 52 56 59 Fibrosis mild 40 37 46 Fibrosis moderate 7 25 26 The increase of bone marrow fibrosis registered after one year (representative also of second year data) resulted not related to patient gender, age >45 years, platelet count >1000 x109/L, Hb <12 g/dL, splenomegaly, previous IFN treatment, PEG Intron dose >50 μg/week. In conclusion, the present study shows that in ET patients a two years PEG Intron treatment, able to induce and to maintain the Hematological Response in the majority of cases, is associated to a decrease of bone marrow cellularity, granulopoiesis, erytropoiesis, MK number, size and ploidy and, moreover, with an increase of MK dystrophy and of bone marrow fibrosis. These preliminary data on bone biopsy and aspirate will be object of a planned centralized reevaluation by a Panel of Pathologists and Clinicians.

Evidence for Reduced Angiogenesis in Bone Marrow of Systemic Sclerosis Patients Immunohistochemetry and Computerized Imagine Analysis.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 5126-5126
Author(s):  
Valentina Carrai ◽  
Renato Alterini ◽  
Riccardo Saccardi ◽  
Irene Miniati ◽  
Luigi Rigacci ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Systemic Sclerosis ◽  
Substantial Reduction ◽  
Bone Marrow Fibrosis ◽  
Bone Marrow Biopsies ◽  
Angiogenic Potential ◽  
Fibrotic Process ◽  
The Mean ◽  
Median Rate ◽  
Marrow Fibrosis

Abstract Abstract 5126 Introdution Systemic sclerosis (SSc) involves the microcirculation, the immune system, and the connective tissue eventually leading to fibrosis. Vascular dysfunction is one of the earliest events in SSc pathogenesis: endothelial damage leads to a dysregulation of angiogenesis and a loss of capillaries. The consequent chronic ischemia provokes a diffuse sufference of the tissues with formation of ulcers. We analyzed BM biopsies in a series of patients undergoing HSCT in our Centre for severe, progressive SSc in order to clarify the association between modification bone marrow angiogenesis and clinical features of this disease. Materials and Methods The main clinical feature of the patients are following: modified Rodnan Skin Score was more than 8 in four patients; nailfold videocapillaroscopy was active in three patients; pulmonary arterial hypertension was upper limit only in two patients; carbon monoxide diffusing capacity was more than 40% in all the patients; high resolution computed tomography showed fibrosis or ground glass lesions in all the patients; topoisomerase I and antinuclear antibody was positive in all the patients. Only one patient showed arrhythmia. Eight SSc bone marrow biopsies were studied, compared with five bone marrow biopsies of non malignant controls. To evaluate angiogenesis, following monoclonal antibodies (MoAb) were used: VEGF, KDR, MMP-9, CD34. Bone marrow fibrosis was evaluated by silver impregnation for reticulum. To identify BM microvessel, anti-CD34 was used. Sections were observed at 400x magnification by two different blinded observers. To calculate the number of vessels, vascular mean area expressed in mm2, vascular percent mean area, perimeter and MVD, a multiparametric, semi-automatic computerized imagine analysis was employed. For each section, we evaluated eight consecutive areas and each area was 16001,92 mm2 (total area was 128015,36 mm2). The VEGF, MMP-9, KDR expression was evaluated as percentage of positive cells in a total of eight consecutive areas at 400x magnification. Results All patients showed a mean cellularity of 40% (SD 5.24, range 30-45%). In four patients bone marrow fibrosis was detected: two patients were classified as I° grade and two as II°grade. The bone marrow biopsy specimens from patients with SSc show a substantial reduction in vascularity. A multiparametric computerized analysis demonstrated significantly reduction of MVD in SSc cases. The mean MVD in SSc bone marrow was 712,63 (SD 392.03, range 124,98-1312,34) whereas in control specimens it was 1364,58 (SD 44.20, range 1312,34-1402,14). The mean number of vessels (p0.004) and percent vascular mean area are lower in sclerosis than controls (p0.0009). A significant increased expression of VEGF was observed. The median VEGF rate was 48,75 (range 30-85%) with expression ≥ 50% in half patients and in two patients with advanced SSc this expression was more than 70%. KDR expression was significantly lower in SSc bone marrow than in controls (p=0,003). The median KDR rate was 13,25% (range 1-40%) and 42% (range 40-45%) in sclerosis and controls respectively. In all cases the expression of MMP-9 was significantly lower than controls (p=0,0009) with median rate of 13,06% (range 1-25%) while median expression in controls was 30% (range 20-40%). Discussion We demonstrated in patient with SSc a reduction of bone marrow vascularity despite the stricking increase of a number of angiogenic factors. VEGF expression in SSc bone marrow is high in all cases with a double median rate compared to controls (48,75% vs 4,25%) while MVD is lower in sclerosis than controls (712,63 vs 1364,58). In regard to these alterations in our study we have observed a reduction of KDR and MMP-9. The overproduction of VEGF can generate a negative feedback to KDR while the low levels of MMP9 found in SSc bone marrow may be due to a hyperproduction of MMP inhibitors contributing to the reduction of bone marrow vascularity. In conclusion, our data demonstrate that in SSc the angiogenic potential of bone marrow is reduced, mirroring the systemic microvascular condition characterised by loss of capillaries and desertification despite the increase of VEGF. The amount of reticular fibers, detected in bone marrow, suggests that the fibrotic process may affect also the bone marrow contributing further to the reduction of angiogenic potential. Disclosures No relevant conflicts of interest to declare.

SNP-A Karyotyping Provides Clinically Relevant Results In Myeloid Hematologic Disorders with Unsuccessful Routine Cytogenetic Testing.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3374-3374
Author(s):  
Kathryn M Guinta ◽  
Hideki Makishima ◽  
Christine L. O'Keefe ◽  
Simon Dujardin ◽  
Ramon V. Tiu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Internal Control ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Prognostic Significance ◽  
Uniparental Disomy ◽  
Treatment Modalities ◽  
Abnormal Karyotype ◽  
Monosomy 7 ◽  
Technical Problems

Abstract Abstract 3374 Metaphase spreads have an established value in the routine diagnostic workup of myeloid malignancies and bone marrow failure disorders. In myelodysplastic syndrome (MDS) abnormal karyotypes play a large role in scoring systems and may greatly impact prognosis, or even predict responsiveness to certain therapies. In aplastic anemia (AA) cytogenetic abnormalities detected by metaphase karyotyping may rule out hypoplastic MDS. In myeloproliferative neoplasia (MPN), an abnormal karyotype may distinguish between reactive or malignant proliferation. Due to technical problems, including specimen quality, viability, hypocellularity or a failure of growth, this routine test may fail to yield conclusive results in some patients. With the advent of SNP-A karyotyping, which only requires extracted DNA, non-informative cases can be resolved. As a cytogenetic test, single nucleotide polymorphism array (SNP-A) analysis can provide an opportunity to improve risk assessment and selection of proper treatment modalities. The advantages of SNP-A include excellent resolution, detection of copy neutral loss of heterozygosity (also known as uniparental disomy or UPD), and perhaps most importantly, the ability to test archived DNA samples, rather than actively dividing cells. However, unlike metaphase cytogenetics, this technology cannot detect subsets of abnormal populations or certain classes of genomic rearrangements, such as balanced translocation, inversion or ring chromosomes. In this study, we examined the prognosis and disease characterization for patients with non-informative cytogenetics (N=144) collected over the last 8 years. SNP-A-based karyotyping has been performed for a representative subset of these patients (N=60) to assess whether this technique could provide clinically relevant information. These patients included patients with MDS (N=20), AA (N=20), AML (N=12) and MDS/MPN (N=3). Bone marrow obtained following induction chemotherapy was excluded. We have detected 27 somatic microdeletions and 33 microduplications (<10Mb) after eliminating germ line copy number variants seen in an internal control cohort (N=1355), publicly available databases or those present in paired non-clonal samples. However, for the purpose of subsequent analysis, somatic microdeletions and duplications were not included, as their prognostic significance has not been validated in large cohorts. (These microalterations may indicate edges of balanced translocations or true clonal pathogenic lesions.) None of these microdeletions were recurrent. Based on these criteria, SNP-A analysis revealed an abnormal karyotype in 14 (23%) patients; 3 with AA and 11 with myeloid disorders. The most common recurrent abnormalities included deletion 5q (N=5) and del7/7q (N=3) but other lesions including13q-, del20, +8p were also seen. Of note we have also detected somatic UPD (regions >25Mb) in 2 cases, including 22q11.23qter and 14q12-q22.1. In 4/60 (7%) a complex karyotype was detected, while 10 had sole lesions (>10Mb). In presumed AA patients, we have identified 2 patients with monosomy 7, prompting a change of diagnosis to MDS and thereby altering their clinical management. In MDS, when cytogenetic prognostic groupings were applied in previously unscored patients, 10/20 had IPSS scores of 3 or greater. The presence of chromosomal abnormalities detected by SNP-At indictated the presence of advanced risk disease and thereby contributed into poorer survival as predicted by IPSS. Disclosures: No relevant conflicts of interest to declare.

Differential Expression of Ribosomal Proteins in Myelodysplastic Syndromes

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1710-1710
Author(s):  
Elizabeth B Rinker ◽  
Julie C. Dueber ◽  
Julianne Qualtieri ◽  
Jason Tedesco ◽  
Begum Erdogan ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Ribosomal Proteins ◽  
Caspase 3 ◽  
Bone Marrow Failure ◽  
Myeloid Cells ◽  
P Value ◽  
Ribosomal Biogenesis ◽  
Erythroid Cells ◽  
Cytoplasmic Staining ◽  
Bone Marrow Failure Syndromes

Abstract Abstract 1710 Introduction: Since the identification of mutations in RPS19 in patients with Diamond-Blackfan anemia (DBA) in 1999,1 proteins in ribosomal biogenesis have been implicated in other congenital bone marrow failure syndromes, including Schwachman-Diamond syndrome (SDS) (SBDS) and X-linked dyskeratosis congenital (DKC1). 2 Collectively known as ribosomopathies, these syndromes share an increased risk of the development of a myelodysplastic syndrome (MDS) and acute myeloid leukemia. MDS is an acquired bone marrow failure syndrome. In the best understood subtype of MDS, 5q- syndrome, deletions of RPS14 in the common deleted region of 5q appear to contribute to the anemia seen in this disease.3 We hypothesize that non-5q minus MDS may also be an acquired ribosomopathy syndrome. Methods: Using published gene expression profiling studies of SDS,4 DBA,5 and MDS,6 the dysregulated ribosomal proteins were compared and 5 ribosomal proteins were identified which represented a union between the three sets of data: RPS10, RPS14, RPS24, RPL14, and RPL36. For practical reasons, we chose to examine RPS14, RPS24, and RPL36. In addition, we selected DKC1, SBDS, CASP3, and Ki67 to further probe the involvement of potential pathways of ribosomal biogenesis, apoptosis, and proliferation. A total of 16 MDS paraffin-embedded bone marrow samples were examined as well as 10 control samples. Immunohistochemical analysis was performed on 4 μm sections from the bone marrow preparations. Staining was performed on the EnVision+ horseradish peroxidase system (DAKO, Carpinteria, CA), using anti-RPS14 (#16683-1-ap, Proteintech, Chicago, IL), anti-RPS24 (#hpa003364, Sigma-Aldrich, St. Louis, MO), RPL36 (#ab74737, Abcam, Cambridge, MA), anti-DKC1 (#nbp1-40097, Novus Biologicals, Littleton, CO), anti-SBDS (#LS-C40532, LifeSpan Biosciences, Seattle,WA), anti-caspase 3 (#mbs440008, MyBioSource, San Diego, CA), and anti-Ki67 (#M7240, Dako, Carpinteria, CA). Expression levels of the proteins were counted in 200 nucleated cells in a blinded fashion by two independent reviewers. A stain index for each sample was determined based upon percentage of positive staining cells per hematologic lineage using upon the counterstain morphology. Unpaired T-test analysis was used to ascertain significance of differences between MDS and normal control samples. Results: Statistically significant changes were found in the ribosomal proteins in at least one cell lineage. Surprisingly, SBDS was found to be expressed in an increased percentage of erythroid cells in MDS (positive cytoplasmic staining in 16% of erythroid cells versus 6% in controls, p value 0.0066). RPL36 was also found to have increased expression in the myeloid lineage in MDS (positive nuclear and cytoplasmic staining in 52% of myeloid cells in MDS versus 33% in controls, p value 0.025) as did RPS24 (positive cytoplasmic staining in 33% of myeloid cells in MDS versus 11% in controls, p value 0.005). RPS14 demonstrated increased expression in myeloid cells (positive cytoplasmic staining in 38% of myeloid cells in MDS versus 18% in controls, p value 0.0042) as well as erythroid cells (positive cytoplasmic staining in 18% of myeloid cells in MDS versus 8% in controls, p value 0.0091), but in fewer megakaryocytes (26% versus 75%, p value 0.038). DKC1 showed increased expression in myeloid nuclei (91% versus 78%, p value 0.043). However, there were no statistically significant differences in expression of caspase 3 or Ki67. Of note, RPS24 is overexpressed while its predicted regulator miR-342 has been shown to be underexpressed in MDS. Similarly, underexpression of miR-10b and miR-150 may correlate with overexpression of DKC1 and correspondingly miRs-378, -140, and -103 with SBDS. Conclusions: Unlike the congenital bone marrow failure syndromes which are associated with loss of function or haploinsufficiency of ribosomal proteins, there appears to be increased expression of ribosomal proteins in the maturing cells of MDS. In contrast to previous studies on CD34+ MDS cells which found decreases in ribosomal protein expression,6 these increases in expression were found on morphologically more mature cells. Several of the ribosomal proteins are predicted to be regulated by miRNAs previously shown to be decreased in MDS.7 There is no overt correspondence with alterations in either apoptosis (measured by caspase 3 expression) or proliferation (Ki67 expression). Disclosures: No relevant conflicts of interest to declare.

Gene Expression Profiling (GEP) of Whole Bone Marrow Biopsies (BMB) in Multiple Myeloma (MM) – Relationship to Plasma-Cell (PC)-Based GEP-Defined Risk and Molecular Subgroups

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3953-3953
Author(s):  
Donald Johann ◽  
Pingping Qu ◽  
Rachael Sexton ◽  
Antje Hoering ◽  
Joshua Epstein ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Conflicts Of Interest ◽  
Risk Groups ◽  
P Value ◽  
Molecular Subgroups ◽  
Gene Score ◽  
Molecular Subgroup ◽  
Bone Marrow Biopsies ◽  
Great Insight ◽  
Prognostic Implications

Abstract Abstract 3953 PC-GEP provides great insight into MM heterogeneity as defined by 7 distinct molecular subgroups, along with clinical correlates and prognosis, as identified by the GEP-70 model. As is now increasingly appreciated in MM, a close interaction exists between PC and the bone marrow micro-environment (ME), which results not only in MM bone disease but also contributes to MM progression and drug resistance. We postulated that such interaction could be revealed by performing GEP both on PC and the bone marrow biopsy (BMB) proper. In addition to having demonstrated that BMB-GEP can attain a normal (NL)-like status in comparison with NL donors with favorable prognostic implications, we are now reporting on paired comparisons of BMB-GEP and PC-GEP toward elucidating whether different molecular subgroups (MOLS) (namely, CD-1, CD-2, MAF, MAF-B, MS, HY, PR) and GEP-70-defined prognostic risk groups (RISK) engage the ME differentially. Toward this end, a recently defined BMB-GEP model of 65 genes, distinguishing NL, MGUS/AMM, MM and MM-CR, was applied to determine whether linkage existed to MOLS and RISK. Indeed, based on mean-based statistics, BMB-65 scores varied among MOLS and RISK (both p=0.013). A negative correlation (R=-0.204) existed between BMB-65 and PC-70 scores (p=0.0014). Further analysis within MOLS revealed significant negative correlations within MF (R=-0.57, p=0.0087) and MS subtypes (R=-0.33, p=0.037). None of the other MOLS showed any significant correlations. This is the first demonstration in patient MM samples that MOLS may engage the ME differentially. The more pronounced inverse correlations of MF and MS subtypes reveal a counter-relationship between RISK and BMB-65 score. This implies that with greater aggressiveness of PC, the BMB appears less NL-like, and is thus more modified to support the malignant PC processes. More detailed analyses will be presented in the context of other GEP and clinical parameters, specifically concerning the prognostic implications of BMB scores at baseline. Correlation between GEP70 score and BMBX 65-gene score within each molecular subgroup Molecular Subgroup N Correlation of GEP70 risk and BMBX 65-gene score P value for testing no correlation CD-1 15 −0.39 0.1494 CD-2 39 −0.22 0.1747 HY 71 −0.15 0.1931 LB 33 0.12 0.4927 MF 20 −0.57 0.0087 MS 40 −0.33 0.0368 PR 25 −0.13 0.55 Overall 243 −0.204 0.001373 Disclosures: No relevant conflicts of interest to declare.

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