scholarly journals Clonal evolution in a patient with aplastic anemia – case report

2015 ◽  
Vol 34 (1-2) ◽  
pp. 45-51
Author(s):  
Melen Brinza ◽  
Cerasela Jardan ◽  
Didona Vasilache ◽  
Camelia Dobrea ◽  
Daniel Coriu
Keyword(s):  
Bone Marrow ◽  
Growth Factors ◽  
Aplastic Anemia ◽  
Young Adulthood ◽  
Clonal Evolution ◽  
Equal Distribution ◽  
Case Presentation ◽  
Men And Women ◽  
Blast Cells ◽  
Serious Disease

Abstract Background: Aplastic anemia (AA) is a rare and serious disease characterized by pancytopenia and hypoplastic bone marrow in the absence of infiltrates/fibrosis. It occurs more frequently in childhood and young adulthood (10-30 years) and with older age (>60 years), with equal distribution among men and women. As hypoplastic myelodysplastic syndromes (hMDS) are also associated with cytopenia and hypocellular marrow,they may be difficult to differentiate from AA. The presence of dysplastic features (others than erythroid) and/or blast cells >5% is essential to distinguish hMDS from AA. Cytogenetic tests may reveal clonal evolution in hMDS. As the two disorders differ greatly in means of management and prognosis, the correct diagnostic is very important. Case presentation: We report the case of a 39 years old female diagnosed in 2005 (at age 29) with aplastic anemia. She received treatment with corticosteroids, Cyclosporine, blood transfusions and growth factors with partial response and no transfusion independency. After 8 years of evolution she developed dysplastic features within the megakaryocytic and granulocytic lineages and an increase in the blast population. The bone marrow slowly became hypercellular. The treatment with cyclosporine and growth factors was stopped.

scholarly journals Nontransplant therapy for bone marrow failure

Hematology ◽  
2016 ◽  
Vol 2016 (1) ◽  
pp. 83-89 ◽  
Author(s):  
Danielle M. Townsley ◽  
Thomas Winkler
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Bone Marrow Failure ◽  
Clonal Evolution ◽  
Single Agent ◽  
Telomere Attrition ◽  
Long Term Follow Up ◽  
Treatment Naïve

Abstract Nontransplant therapeutic options for acquired and constitutional aplastic anemia have significantly expanded during the last 5 years. In the future, transplant may be required less frequently. That trilineage hematologic responses could be achieved with the single agent eltrombopag in refractory aplastic anemia promotes new interest in growth factors after years of failed trials using other growth factor agents. Preliminary results adding eltrombopag to immunosuppressive therapy are promising, but long-term follow-up data evaluating clonal evolution rates are required before promoting its standard use in treatment-naive disease. Danazol, which is traditionally less preferred for treating cytopenias, is capable of preventing telomere attrition associated with hematologic responses in constitutional bone marrow failure resulting from telomere disease.

Pediatric Aplastic Anemia and Refractory Cytopenia of Childhood: A Retrospective Single Institutional Analysis to Determine Outcomes and Histomorphological Predictors

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4382-4382
Author(s):  
Craig Forester ◽  
Sarah E Sartain ◽  
Dongjing Guo ◽  
Marian Harris ◽  
Olga Weinberg ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Treatment Failure ◽  
Aplastic Anemia ◽  
Disease Progression ◽  
Clinical Diagnosis ◽  
Prognostic Value ◽  
Bone Marrow Failure ◽  
Clonal Evolution ◽  
Bone Marrow Failure Syndromes ◽  
Consensus Diagnosis

Abstract Introduction Pediatric acquired aplastic anemia (AA) is a hypocellular bone marrow condition that is often difficult to distinguish from inherited bone marrow failure syndromes (IBMFS) and hypoplastic refractory cytopenia of childhood (RCC). RCC is a provisional entity for which the clinical implications are still under investigation. Historically, patients with RCC have been classified as AA. In this study, we sought to assess the intra-observer reproducibility and prognostic value of the histological criteria for RCC and AA as defined in the WHO classification. Specifically, we evaluated if RCC is an independent prognostic factor of overall survival (OS) and event-free survival (EFS), including treatment failure, disease progression and clonal evolution. Methods We performed a retrospective analysis of 149 AA patients seen at a single center between 1976-2010. Patients that met clinical or molecular diagnostic criteria for inherited bone marrow failure syndromes were excluded. We evaluated 5-year EFS, OS, and response rate to immunosuppressive therapy (IST); outcomes after matched HLA-identical related donor transplant (MRD) and matched unrelated donor (MUD) stem cell transplantation, as well as toxicities and clonal evolution. Of the 149 eligible individuals, 72 had diagnostic pathology material available for review. A double-blinded analysis of bone marrow aspirate and biopsy slides was undertaken by 3 pediatric hematopathologists. Each pathologist reviewed the slides of the entire study set and independently made a determination of the diagnosis. In cases where one or more differed in their assessment, the three conferred with an additional senior pediatric hematopathologist and all four reached agreement on a ‘consensus diagnosis’. To describe the degree of concordance between the assessments of the pathologists, a kappa coefficient was calculated. A logistic regression model was used to identify factors prognostic of treatment failure. Survival curves were generated using the methods of Kaplan and Meier. Results The 149 patients were classified as moderate aplastic anemia (MAA) (n=58), severe aplastic anemia (SAA) (n=50) and very severe aplastic anemia (vSAA) (n=41). Ninety-one patients received IST, 50 underwent a MRD HSCT, and 8 were observed without treatment. The concordance between pathologists in the assessment of the diagnosis of AA or RCC was modest, but ultimately a consensus between the pathologists was reached. The overall EFS and OS were 50.8% and 73.1% for all patients. The 5-year OS and 5-year EFS for all patients receiving IST were 87.8% and 51.5%, respectively, with a failure to respond to IST in 48%. Patients with vSAA had worse 5-year OS compared to SAA and MAA patients, respectively (p=0.02). 5-year OS was comparable at 83.6%±7.2 and 85.3%±5.2 for patients receiving a MUD or MRD respectively. Within AA patients receiving IST, none of the following factors had prognostic value to predict failure of treatment: consensus diagnosis of RCC, macrocytosis (MCV > 100), or HgF% > 4%. An MCV >100and HgF% >4% likewise were not associated with consensus diagnosis of RCC. However, a significantly higher hemoglobin (Hgb) and absolute reticulocyte count was found in patients with RCC compared to AA. Surprisingly the diagnosis of RCC was associated with a trend towards improved 5- year OS (84.9% versus 72.5%) and 5-year EFS (71.3% versus 52.5%) compared to AA. Five patients (~3%) of the analytical cohort all with the clinical diagnosis of MAA experienced clonal evolution or disease progression. Interestingly, the clinical diagnosis of vSAA significantly correlated with the histologic appearance of AA, whereas SAA and MAA corresponded with RCC, strongly suggesting a correlation between lower disease severity as determined by peripheral counts and/or marrow cellularity and the diagnosis of RCC. Conclusion The EFS and OS of the IST group in this single institution study is consistent with larger series. Our data indicate a low rate of clonal evolution in pediatric AA that in this series was associated exclusively with MAA. Most important, our findings suggest that the histologic diagnosis of RCC, reached by consensus among four pediatric hematopathologists does not predict disease outcome in this retrospective series of AA. These data indicate the need for larger prospective studies to determine the clinical significance of the classification. Disclosures Sartain: Hemostasis and Thrombosis Research Society: Research Funding.

scholarly journals Morphology in patients with severe aplastic anemia treated with antilymphocyte globulin

Blood ◽  
1992 ◽  
Vol 80 (2) ◽  
pp. 337-345 ◽  
Author(s):  
A Tichelli ◽  
A Gratwohl ◽  
C Nissen ◽  
E Signer ◽  
C Stebler Gysi ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Peripheral Blood ◽  
Clonal Evolution ◽  
Cumulative Risk ◽  
Fetal Hemoglobin ◽  
Severe Aplastic Anemia ◽  
Antilymphocyte Globulin ◽  
Probability Of Survival ◽  
Patients At Risk

One hundred and seventeen patients with severe aplastic anemia (SAA) were treated at our institution between 1976 and 1990 with antilymphocyte globulin (ALG) therapy. Seventy-nine (68%) are alive and probability of survival at 14 years, according to Kaplan and Meier, is 62% +/- 12%. Twenty-six patients developed a late clonal complication: 11 had a myelodysplastic syndrome (MDS) and 17 had paroxysmal nocturnal hemoglobinuria (PNH); two patients had both. The cumulative risk at 10 years is 42%. The development of MDS/PNH after SAA directly affects survival. The probability of being alive at 14 years is 81% +/- 10% for patients with stable disease and 36% +/- 13% for those with clonal evolution (P = .001). To look for predictive signs, we reevaluated peripheral blood and bone marrow cytomorphology at presentation, during regeneration, and in remission. We examined the peripheral blood values for hemoglobin, reticulocytes, granulocytes, thrombocytes, mean corpuscular volume (MCV), and fetal hemoglobin, as well as bone marrow for cellularity, erythropoiesis, myelopoiesis, and megakaryopoiesis. ALG therapy induces slow and incomplete recovery. Although in “remission,” ALG patients have lower hemoglobin values, higher reticulocyte counts, lower granulocyte and platelet values, and a higher MCV and fetal hemoglobin than normal controls. They retain a reduced number of megakaryocytes and a persistence of atypical monocytes in bone marrow morphology as stigmata of their disease. Patients with late clonal complications show distinct morphologic abnormalities: patients with PNH have higher MCVs, higher granulocyte and reticulocyte counts, and more dyserythropoiesis at diagnosis and a lower hemoglobin with an increased proportion of erythroblasts in the bone marrow in “remission.” Patients who later developed MDS are not different from the total patient population at diagnosis. After therapy, these patients are characterized by the presence of ring sideroblasts and atypical monocytes during regeneration and by a persistent increase in MCV, a higher fetal hemoglobin, lower granulocyte values, and megakaryocytic dysplasia during “remission.” Thus, routine morphologic follow-up examination of blood and bone marrow can discover patients at risk for late hematologic complications after ALG therapy.

Telomere Shortening Promotes Chromosomal Instability and Predicts Malignant Clonal Evolution in Aplastic Anemia.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3208-3208
Author(s):  
Rodrigo T. Calado ◽  
James N Cooper ◽  
Phillip Scheinberg ◽  
Colin Wu ◽  
Marco A Zago ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Telomere Length ◽  
Malignant Transformation ◽  
Chromosomal Instability ◽  
Bone Marrow Cells ◽  
Clonal Evolution ◽  
Monosomy 7 ◽  
Marrow Cells

Abstract Abstract 3208 Poster Board III-145 In murine models, telomere erosion promotes chromosomal instability via breakage-fusion-bridge cycles, contributing to the early stages of tumorigenesis. However, direct evidence that short telomeres predispose to cancer development in humans is lacking. In acquired aplastic anemia, evolution to malignant clonal disorders is a major complication after immunosuppressive therapy, affecting up to 15 percent of patients at 10 years. We investigated whether telomere length measured at diagnosis predicted clonal evolution in these patients. Telomere length was measured from DNA extracted from peripheral blood leukocytes collected at disease presentation in 183 consecutive patients enrolled in successive clinical trials for immunosuppressive regimen as first line therapy for severe aplastic anemia at the Clinical Research Center, National Institutes of Health (ClinicalTrials.gov identifier numbers, NCT00001964, NCT00260689, and NCT00061360) and 164 healthy volunteers. Leukocyte telomere length of aplastic anemia patients at diagnosis was in the normal range and was not shorter than in healthy controls (ANOVA-F test). Telomere length was corrected for age and patients were separated into two groups: patients with short telomeres (in the lowest quartile) and long telomeres (other quartiles). Telomere length was a critical and independent predictive biomarker for evolution to myelodysplastic syndrome, especially monosomy 7, and acute myeloid leukemia (AML) in patients with acquired aplastic anemia (Multivariate Cox Proportional Hazard Model, P=0.006). Patients with short telomeres had six-fold higher probability to develop clonal malignant disease than did patients with longer telomeres. Bone marrow cells of aplastic patients were cultured in vitro for short term in the presence of cytokines and high-dose granulocyte-colony stimulating factor (G-CSF) and cells of patients with short telomeres (n=5) showed increased telomere-free chromosomal ends in comparison to cells of patients with long telomeres (n=6), by fluorescence in situ hybridization (FISH; P<0.0001). Spectral karyotyping (SKY) revealed that cultured bone marrow cells of patients with short telomeres exhibited aneuploidy and translocations, including Robertsonian translocations, which were not found in cells of patients with long telomeres. Bone marrow cells at diagnosis were further evaluated for the presence of monosomy 7 cells using interphase FISH in 73 patients. Telomere length inversely correlated with the frequency of monosomy 7 cells: the shortest the telomeres, the highest the percentage of aneuploid cells at diagnosis (Pearson r=-0.5110; P=0.0009). We further employed bone marrow cells of clinically healthy individuals carrying loss-of-function telomerase mutations and with extremely short telomeres (n=5) as a model for telomere dysfunction in hematopoietic cells in the absence of human disease. In vitro culture of these cells yielded aberrant karyotypes by SKY, including translocations and aneuploidy, and end-to-end chromosomal fusions by FISH. These results indicate that telomere length at diagnosis predicts evolution to myelodysplasia and leukemia in patients with acquired aplastic anemia treated with immunosuppression. Our findings support the hypothesis that short and dysfunctional telomeres restrain stem cell proliferation and predispose for malignant transformation by selecting stem cells that are prone to chromosomal instability. This is the first prospective study to demonstrate that short telomeres in human hematopoietic cells promote chromosomal instability in vitro and predispose to malignant transformation in humans. Disclosures Cooper: NIH-Pfizer: Research Funding.

Hematopoietic Growth Factors In Aplastic Anemia Patients Treated with Immunosuppressive Therapy: Systematic Review and Meta-Analysis - 2010 Update.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1164-1164
Author(s):  
Ronit Gurion ◽  
Anat Gafter-Gvili ◽  
Liat Vidal ◽  
Mical Paul ◽  
Isaac Ben-Bassat ◽  
...  
Keyword(s):  
Systematic Review ◽  
Growth Factors ◽  
Aplastic Anemia ◽  
Immunosuppressive Therapy ◽  
Clonal Evolution ◽  
Meta Analysis ◽  
Cochrane Library ◽  
Hematopoietic Growth Factors ◽  
Hematologic Response ◽  
All Cause Mortality

Abstract Abstract 1164 Background: Immunosuppressive therapy (IST) is the treatment for patients with severe aplastic anemia (SAA) not eligible for transplantation. It is controversial whether there is a role for hematopoietic growth factors (HGF) as an adjunct to IST in these patients. Objectives: A meta-analysis evaluating the role of HGF in this setting was published by our group in 2009. Since then, results of the largest conducted clinical trial by the Aplastic Anemia Working Party of the EBMT have been reported. We therefore updated our meta-analysis in order to evaluate if in 2010 there is still a role for the addition of HGF to IST in patients with SAA. Methods: Systematic review and meta-analysis of randomized controlled trials comparing treatment with IST and HGF to IST alone in patients with SAA. An updated search in The Cochrane Library, MEDLINE, conference proceedings and references was conducted in July 2010. Two reviewers independently assessed the quality of the trials and extracted data. Outcomes assessed were: all-cause mortality, overall hematologic response, infections and clonal evolution (transformation to myelodysplastic syndrome or acute leukemia). Relative risks (RR) with 95% confidence intervals (CIs) were estimated and pooled. Results: Our search yielded 7 trials, randomizing 619 patients, including the 205 patients included in the EBMT trial recently published. Trials were conducted between the years 1991 and 2008. The IST regimen for most trials consisted of anti-thymocyte globulin, cyclosporine and steroids. The HGF in 6 trials was G-SCF and in 1 trial GM-CSF and erythropoietin. The addition of HGF to IST, compared with IST alone yielded no difference in all cause mortality at 100 days (RR 1.33, 95% CI 0.56–3.18) and at 5 years (RR 0.91, 95% CI 0.64–1.30, Fig.1). There was no difference in overall hematologic response at 12 months between the two arms (RR 1.16, 95% CI 0.91–1.47). There was no increase in the incidence of clonal evolution in the HGF arm compared to the control (RR 1.45, 95% CI 0.42–5.07). In addition there was no difference in the number of infections between both arms (RR 0.98, 95%CI 0.82–1.17). Conclusions: The addition of HGF to IST in SAA does not influence all-cause mortality, long term response, or the incidence of infections. The cumulative data in our updated meta-analysis is consistent with the results of our previous report. Therefore, HGFs should not be recommended routinely as an adjunct to IST for patients with SAA. Disclosures: Shpilberg: Roche: Consultancy, Honoraria.

The Usefulness of CD71 Expression by Flow Cytometry in the Diagnosis of Acute Leukemia.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2533-2533 ◽  
Author(s):  
Qian Liu ◽  
Mangju Wang ◽  
Yang Hu ◽  
Haizhou Xing ◽  
Xue Chen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Leukemia ◽  
Clonal Evolution ◽  
Expression Level ◽  
Leukemic Cells ◽  
Bone Marrow Blast ◽  
Blast Cells ◽  
The Mean ◽  
Acute Megakaryocytic Leukemia ◽  
Normal Controls

Abstract Abstract 2533 CD71 (transferring receptor 1) is an integral membrane glycoprotein that plays an important role in cellular uptake of iron. It is well known as a marker for cell proliferation and activation. Although all proliferating cells in hematopoietic system express CD71, however, CD71 has been considered as a useful erythroid-associated antigen. The expression proportion on nucleated red blood cells was significantly higher than other cells, approximately 80% of all CD71 positive cells were of CD71 positive erythroid cells in normal bone marrow. CD71 was usually considered as the representative marker for differentiating erythroblasts and diagnosing acute erythroid leukemia (AEL) by flow cytometry. At the ISAC 2000 Congress, most experts agreed that at least one or more B, T, myeloid, erythroid and megakaryocytic reagents should be included in the essential panel. The reagents recommended for erythroid cells included CD36, CD71 and glycophorin A (GlyA). However, there was no agreement on how to choose and group these antibodies. In the practical analysis of immune phenotypes of leukemic cells we noted that no CD71 expression was detected on blasts of some cases of AEL with typical morphological and cytochemical findings, but other types of acute myeloblastic leukemia (AML) cells may express CD71. Thus, we speculated that CD71 expression may associate with the abnormal antigen expression resulting from hematopoietic disorders. In this study, we evaluated CD71 expression on different acute leukemia cells in association with a variety of other antibodies. In this study we aimed to define CD71 as a flow cytometric marker for the diagnosis of acute leukemia. Bone marrow samples were collected from 82 newly diagnosed acute leukemia patients as well as 13 normal controls. The diagnosis were made according to the WHO 2008 diagnostic criteria. All 6 cases of AEL were erythroid/myeloid subtype (acute erythroid/myeloid leukemia, M6a). The samples were then analyzed using a four-color flow cytometer with antibody panels against a variety of lymphoid, myelomonocytic, erythroid and megakaryocytic antigens. The antibodies included anti-CD3, CD7, CD10, CD11b, CD13, CD14, CD15, CD16, CD19, CD20, CD33, CD34, CD45, CD56, CD61, CD64, CD71, CD117, GlyA, HLA-DR, IgG, IgM, MPO. Subpopulations of bone marrow cells were gated based on CD45 intensities and side scatter (SSC) value to further analyze the expression of antigens in different cell populations. Positive CD71 expression were identified on bone marrow blast cells of 41 (50%) acute leukemia patients and 9 (69.23%) normal controls. The mean expression level on normal controls was 35.99±19.06%. The mean CD71 expression level on blasts of AML with blasts differentiation at early stage of myelopoiesis (including FAB-M0/M1/M2/M4) was significantly higher than AML with partial differentiation of leukemic cells (FAB-M3/M5) and acuteB lymphoblastic leukemia (B-ALL) (p<0.05), with the mean expression level of 38.78±26.65%, 13.25±8.75% and 10.12±11.65%, respectively, and the latter two lower than normal controls (p<0.05). The percentage of CD71 expression level on blasts of acute megakaryocytic leukemia (FAB-M7) was 80.16±8.23%, significantly higher than normal controls, partial differentiation of leukemic cells (FAB-M3/M5), and B-ALL (p<0.05). The percentage of CD71 expression level on blasts of mixed lineage leukemia was 49.66±22.69%, significantly higher than B-ALL (p<0.05). Positive CD71 expression was found on bone marrow blast cells of 4 (66.67%) AEL cases, with the mean level percentage of 25.68±11.63% that was significantly lower than acute megakaryocytic leukemia (FAB-M7) (p<0.05) and was indifferent from normal controls and other types of acute leukemia. Using CD71 expression levels, we identified different abnormal cell clones simultaneously existing within bone marrow of 2 patients of AML with maturation (FAB-M2) and AEL, implicating the clonal evolution process from normal blasts to leukemic cells. CD71 is an important marker for diagnosing acute leukemia, and is useful for distinguishing the differentiation stages of AML. However, CD71 may not be the specific diagnostic marker for AEL. CD71 is also valuable for the observation of clonal evolution process of acute leukemia, which may be informative to the etiology of leukemia. Disclosures: No relevant conflicts of interest to declare.

Clonal Evolution In Aplastic Anemia Is Driven By Chromosomal Instability Rather Than Mutations In Myeloid Malignancy Candidate Gene

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 802-802 ◽  
Author(s):  
Bogdan Dumitriu ◽  
Xingmin Feng ◽  
Yasutaka Ueda ◽  
Sachiko Kajigaya ◽  
Danielle Townsley ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Telomere Length ◽  
Tissue Regeneration ◽  
Chromosomal Instability ◽  
Clonal Evolution ◽  
Heterozygous Mutation ◽  
Short Telomere ◽  
Monosomy 7 ◽  
Telomere Attrition

Abstract The pathophysiology of human aplastic anemia (AA) is immune-mediated destruction of bone marrow stem and progenitor cells. Most patients respond to immunosuppressive therapies (IST), which markedly improved survival in this disease. However, a minority of patients undergoes transformation to malignant hematologic disease, myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML), usually accompanied by the cytogenetic abnormality monosomy 7. Clonal evolution in AA confers a poor prognosis in the clinic but is an opportunity to assess early events in oncogenesis in the setting of inflammation and tissue regeneration. We previously reported that low mean telomere length of leukocytes at the time of diagnosis of AA (Scheinberg et al., JAMA 2010) was associated with increased risk of progression to MDS. In the current study, we directly compared acquired mutations in candidate genes and chromosomal instability, as measured by telomere length, in a cohort of AA patients that had progressed to MDS. Thirteen AA patients who developed monosomy 7 were compared with 30 AA patients who had received similar treatments but did not progress to MDS. Leukocytes telomere content was measured by qPCR in samples obtained at different time points from the diagnosis of AA and chromosome-specific telomere length was assessed by single telomere length assay (STELA) for Xp, Yp, 12q, and 17p. In the AA patients who had evolved to MDS and AML, analysis of acquired mutations in myeloid-specific genes was performed by comparison with control “germline” DNA from purified CD3 lymphocytes by exome sequencing. Cells from AA patients with clonal evolution showed marked progressive telomere attrition, 419 bp/year during the period preceding development of monosomy 7. Telomere attrition was progressive from the time of diagnosis of AA. By STELA, accumulation of very short telomere fragments was apparent at 6 months after IST. In contrast, for the AA control group, patients whose disease was stable, telomere attrition was not accelerated in serial qPCR determinations, nor was there increased accumulation of short telomere fragments by STELA. A similar pattern of increased telomere attrition was reproduced in vitro by cultivation of bone marrow cells obtained six months after IST in all AA patients who developed MDS, while none of the AA control bone marrow cultured cells developed shorter telomeres. We examined bone marrow myeloid cells at the time of monosomy 7 for acquired mutations in 125 candidate genes reported to be recurrently mutated in AML and MDS. Exome sequencing was performed using Agilent SureSelect Target Enrichment System. The raw reads were mapped to UCSC Human Genome hg19 by BWA software with default setting. With an average of 111-fold coverage on selected exon regions, somatic mutations were identified between paired samples using SAMtools and Shimmer software for SNP detection. Acquired mutations in myeloid cells were found in two cases. One patient had a heterozygous mutation in DNMT3A (K829T) present since diagnosis of AA. The other patient also had a heterozygous mutation in DNMT3A (P904S) as well as mutations in DOTL1, ASXL1, SETBP1, and STAT3. All these mutations were identifiable after IST as neutrophils recovered. Despite the presence of multiple mutations, this patient had shown a good hematologic response to IST; evolution was manifest as recurrent pancytopenia and stable marrow myeloblasts at about 5% for over 2 years after first detection of monosomy 7. The remaining 11 patients, all of whom lacked candidate gene mutations, had progressive increase in bone marrow myeloblast numbers; the only other three survivors in this cohort had received hematopoietic stem cell transplant. In conclusion, telomere shortening rather than accumulation of point mutations in hematopoietic cells preceded aneuploidy and malignant transformation at an early stage of oncogenesis in this group of patients. These results from AA may be generalizable to other cancers arising in the setting of inflammation and tissue regeneration in other organs. Identification of critically short telomeres before the development of cytogenetic abnormalities may allow for improved management of patients at risk of clonal evolution, and pharmacologic strategies to increase telomerase activity might mitigate the risk of cancer in these settings Disclosures: No relevant conflicts of interest to declare.

Germline and Somatic Genetic Characterization of Shwachman-Diamond Syndrome

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2681-2681
Author(s):  
Kasiani C. Myers ◽  
Adam S. Nelson ◽  
Brian Sheehan ◽  
Maggie Malsch ◽  
Gwendolyn Towers ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Diagnostic Criteria ◽  
Congenital Anomalies ◽  
Clonal Evolution ◽  
Severe Aplastic Anemia ◽  
Splice Donor ◽  
Shwachman Diamond Syndrome ◽  
Intron 2

Abstract The North American Shwachman-Diamond Syndrome Registry (SDSR) opened in 2008 to improve our understanding of the natural history of Shwachman-Diamond Syndrome (SDS), improve medical outcomes, and facilitate research. The diagnosis of SDS was defined by either biallelic SBDS gene mutations or by the clinical combination of exocrine pancreatic dysfunction with bone marrow failure. Median age of study subjects is 11.2 years (range, 0.6-52.8). SBDS genetic reports were available for 168 subjects. Eighty-one had biallelic SBDS mutations, while 48 individuals lacked SBDS mutations. Ongoing characterization of SBDS mutation-negative individuals has identified subgroups of SDS individuals meeting clinical diagnostic criteria as outlined above, as well as a more heterogenous subgroup with features of SDS but in whom a clinical diagnosis could not be confirmed by classic diagnostic criteria. Amongst those with biallelic SBDS mutations, cytopenias were noted in all but 1 subject. Intermittent neutropenia was noted in 97% (n=70/72) and was the most frequent hematologic abnormality. Anemia was noted in 53% (n=39/73) and thrombocytopenia in 64% (n=48/75). Congenital anomalies were seen in 50% (n=39/78). The mutational spectrum of SBDS was explored. In 80 of 81 patients harboring biallelic SBDS mutations, the c.258+2 T>C intron 2 splice donor mutation was found in at least one of the two mutant SBDS alleles. In these cases, the mutation spectrum of the second mutant SBDS allele included missense mutations, splice site mutations, and truncating mutations. The one patient lacking the intron 2 splice donor mutation had a c.183_184delTAinsCT mutation together with a c.523 C>T mutation in SBDS confirmed to be in trans. This results in the combination of a truncating p.Lys62X mutation with a p.R175W missense mutation at a conserved residue predicted to be deleterious. This patient presented with neonatal severe aplastic anemia requiring platelet and red cell transfusions with neutrophil counts of 0-200 unresponsive to G-CSF. The marrow showed irregular islands of cartilage surrounded by osteoid consistent with a disorder of bone formation. A low level of SBDS protein is expressed by the c.258+2T>C variant, so the absence of this hypomorphic allele may have contributed to this exceptionally severe phenotype. Bone marrow reports were available for 67 subjects with biallelic SBDSmutations. Marrow hypocellularity was noted in 79% (n=49/62). Mild morphologic marrow dysplasia was observed in 58% (n=35/60). Clonal abnormalities developed in 36%. The most common clonal abnormality was del20q in 16% (n=10/64). Isochromosome 7 was noted in 2% (n=1/64). Three individuals developed AML at ages 19.5, 38, and 39 years. Eleven (14%) have undergone hematopoietic stem cell transplantation (HSCT), 10 for MDS or AML and 1 for severe aplastic anemia. Given the frequency of del20q clones in SDS, we studied clinical features of the 10 SDSR subjects with del20q clones. Median age of this group was 17 years (range, 10.8-29.3). Congenital anomalies were noted in 80% (n=8/10). Frequency of cytopenias was similar to that of non-del20q subjects, with neutropenia, anemia and thrombocytopenia seen in 90%, 50%, and 80% respectively. Bone marrow pathology reports were available for 9 subjects. All were noted to have hypocellular marrows as well as mild marrow dysplasia. In many patients, the clone was persistent or grew over time, with longest duration of 14.6 years. Progression to MDS was reported in 3 subjects who initially had an isolated del20q clone. Two developed an additional loss of chromosome 7. Progressive marrow dysplasia and falling blood counts were seen in two subjects. All three were treated with HSCT at 5.4, 5.7 and 7 years. Deletion of 20q in SDS has been hypothesized to result in a milder hematologic phenotype due to compensatory deletion of eIF6. eIF6 binds to the nascent 60S ribosomal subunit and sterically inhibits joining of the 60S to the 40S ribosomal subunits. SBDS functions to facilitate release of eIF6 thus promoting assembly of the mature 80S ribosome. These data from the SDSR suggest that SDS patients with del20q clones remain at risk for clonal evolution. Higher patient numbers are needed to quantitate risk of MDS in SDS patients with del20q. The SDSR continues to expand and mature as a resource for biological and clinical studies in this rare disorder advancing our understanding of marrow failure and clonal evolution. Disclosures Davies: Novartis: Honoraria. Dale:Amgen: Consultancy, Honoraria, Research Funding.

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