scholarly journals Landscape Of Genetic Lesions In 944 Patients With Myelodysplastic Syndromes

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 521-521 ◽  
Author(s):  
Yasunobu Nagata ◽  
Vera Grossmann ◽  
Yusuke Okuno ◽  
Ulrike Bacher ◽  
Genta Nagae ◽  
...  
Keyword(s):  
Myelodysplastic Syndromes ◽  
Cox Regression ◽  
Molecular Model ◽  
Clonal Evolution ◽  
Univariate Analysis ◽  
Gene Mutations ◽  
Risk Groups ◽  
Employment Equity ◽  
Linear Predictor ◽  
Equity Ownership

Abstract Background Myelodysplastic syndromes (MDS) are a heterogeneous group of myeloid neoplasms characterized by varying degrees of cytopenias and a predisposition to acute myeloid leukemia (AML). With conspicuous clinical and biological heterogeneity in MDS, an optimized choice of treatment based on accurate diagnosis and risk stratification in individual patients is central to the current therapeutic strategy. Diagnosis and prognostication in patients with myelodysplastic syndromes (MDS) may be improved by high-throughput mutation/copy number profiling. Methods A total of 944 patients with various MDS subtypes were screened for gene mutations and deletions in 104 known/putative genes relevant to MDS using targeted deep-sequencing and/or array-based genomic hybridization. Impact of genetic lesions on overall survival (OS) was investigated by univariate analysis and a conventional Cox regression, in which the Least Absolute Shrinkage and Selection Operator (lasso) was used for selecting variables. The linear predictor from the Cox regression was then used to assign the patients into discrete risk groups. Prognostic models were constructed in a training set (n=611) and confirmed using an independent validation cohort (n=175). Results After excluding sequencing/mapping errors and known or possible polymorphisms, a total of 2,764 single nucleotide variants (SNVs) and insertions/deletions (indels) were called in 96 genes as high-probability somatic changes. A total of 47 genes were considered as statistically significantly mutated (p<0.01). Only 6 genes (TET2, SF3B1, ASXL1, SRSF2, DNMT3A, and RUNX1) were mutated in >10% of the cases. Less common mutations (2−10%) involved U2AF1, ZRSR2, STAG2, TP53, EZH2, CBL, JAK2, BCOR, IDH2, NRAS, MPL, NF1, ATM, IDH1, KRAS, PHF6, BRCC3, ETV6, and LAMB4. Intratumoral heterogeneity was evident in as many as 456 cases (48.3%), even though the small number of gene mutations available for evaluation was thought substantially to underestimate the real frequency. The number of observed intratumoral subpopulations tended to correlate with the number of detected mutations and therefore, advanced WHO subtypes and risk groups with poorer prognosis. Mean variant allele frequencies (VAFs) showed significant variations among major gene targets, suggesting the presence of clonogenic hierarchy among these common mutations during clonal evolution in MDS. The impact of these genetic lesions on clinical outcomes was initially investigated in 875 patients. In univariate analysis, 25 out of 48 genes tested significantly affected overall survival negatively (P<0.05), and only SF3B1mutations were associated with a significantly better clinical outcome. Next, to evaluate the combined effect of these multiple gene mutations/deletions, together with common clinical/cytogenetic variables used for IPSS-R, OS was modeled by a conventional Cox regression. A total of 14 genes, together with age, gender, white blood cell counts, hemoglobin, platelet counts, cytogenetic score in IPSS-R, were finally selected for the Cox regression in a proportional hazard model and based on the linear predictor of the regression model, we constructed a prognostic model (novel molecular model), in which patients were classified into 4 risk groups showing significantly different OS (“low”, “intermediate”, “high”, and “very high risk”) with 3-year survival of 95.2%, 69.3%, 32.8%, and 5.3%, respectively (P<0.001). These results demonstrated that the mutation/deletion status of a set of genes could be used as variables independent of clinical parameters to build a clinically relevant prognostic score. When applied to the validation cohort, the novel molecular model was even shown to outperform the IPSS-R. Conclusions Large-scale genetic and molecular profiling by cytogenetics, NGS and array-CGH not only provided novel insights into the pathogenesis and clonal evolution of MDS, but also helped to develop a powerful prognostic model based on gene mutations and other clinical variables that could be used for risk prediction. Molecular profiling of multiple target genes in MDS is feasible and provides an invaluable tool for improved diagnosis, biologic subclassification and especially prognostication for patients with MDS. Disclosures: Grossmann: MLL Munich Leukemia Laboratory: Employment. Bacher:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Alpermann:MLL Munich Leukemia Laboratory: Employment. Roller:MLL Munich Leukemia Laboratory: Employment. Nadarajah:MLL Munich Leukemia Laboratory: Employment. Kohlmann:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

scholarly journals Impact of Amotosalen/UVA Pathogen Inactivated Platelet Components on Outcomes after Allogeneic Hematopoetic Stem Cell Transplantation: A Single Center Analysis

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1167-1167
Author(s):  
Andreas S. Buser ◽  
Laura Infanti ◽  
Andreas Holbro ◽  
Joerg Halter ◽  
Sabine Gerull ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cox Regression ◽  
Univariate Analysis ◽  
Conditioning Regimen ◽  
Disease Status ◽  
Risk Scores ◽  
Employment Equity ◽  
Stem Cell Source ◽  
Equity Ownership ◽  
The Impact

Background: Platelet component (PC) transfusion is required for allogeneic hematopoietic stem cell transplantation (HCT) recipients. Contamination with infectious pathogens (bacteria, viruses, or protozoa) and T-cells is a risk factor for transfusion-transmitted infection (TTI) and transfusion associated graft-versus-host disease (TA-GVHD). Pathogen inactivation (PI) treatment of PC with amotosalen-UVA (PI-PC, INTERCEPT Blood System, Cerus Corp) in platelet additive solution (PAS) without bacterial screening, gamma irradiation, CMV serology, and with 7-day storage has been the standard of care in Switzerland since 2011 to manage risk of TTI and TA-GVHD. PI-PC have replaced conventional PC (C-PC) prepared in PAS with gamma irradiation and 5 day storage. We previously reported platelet usage in two consecutive five year periods at the University Hospital of Basel. Mean PI-PC dose was higher (3.0 vs. 2.8 x 1011, p=0.001) and mean storage duration longer (4.2 vs. 3.4 days: p=0.001) than with C-PC. PC expiration wastage was reduced with 7-day PI-PC storage vs. 5-day storage (1.5% vs. 8.7%). For HCT recipients, days of PC support; PC use per patient; and RBC use per patient were similar, despite 24.3% lower corrected count increments (CCI) with PI-PC. Now, we report the impact of these observations on treatment related mortality (TRM) and overall survival (OS) 100 days after HCT. Patients and Methods: A two-period retrospective cohort study was conducted to evaluate PI-PC impact on outcomes of consecutive first allogeneic HCT recipients from January 2006 to December 2010 (Period 1, P1), when gamma-irradiated apheresis C-PC were used, and Period 2 (P2) from January 2011 to December 2017, when apheresis and whole blood-derived PI-PC were used. The review utilized 100-day OS and 100-day TRM to determine the impact of PI-PC on HCT outcomes. Descriptive statistics were used for continuous variables and log-rank analysis for survival outcomes. Univariate analysis was performed using Pearson χ2 statistics. Multivariate Cox regression modelling analyses included: PC period (P1, P2), donor match (HLA identical/twin, matched related, matched unrelated), disease state (early, intermediate, late), and conditioning regimen (reduced intensity, myeloablative) with TRM as the outcome. This was an IRB approved single-center analysis. Results: In P1 and P2, 256 and 557 consecutive first-time allogeneic HCT recipients were included, respectively. By univariate analysis, the distribution of European Group for Bone Marrow Transplantation (EBMT) risk scores (grouped 0-2, 3-4, 5-7) and mean patient age were higher during P2 (p = 0.001 and p <0.001, respectively). Primary disease status (p = 0.039); stem cell source (p <0.001); GVHD prophylaxis with ATG (p <0.001); total body irradiation (p <0.001); and conditioning regimen (p <0.001) were different between P1 and P2. Donor match (p=0.084) and disease status (p = 0.628) were similar in P1 and P2. TRM at day 100 post HCT was significantly less (31/557, 5.5%) for PI-PC recipients in P2 vs. C-PC recipients in P1 (37/256, 14.5%, p<0.001). Overall proportion of survivors at day 100 post HCT was significantly greater for PI-PC recipients (507/557, 91.0 %) compared to C-PC recipients (209/256, 81.6%, p <0.001). By multivariate Cox regression analysis, P2 with PI-PC component support was associated with improved TRM (p = 0.001; adjusted hazard ratio 0.433; 95% confidence interval: 0.262, 0.716). Donor match (p = 0.019), disease state (p = 0.022), and myeloablative conditioning (p = 0.034) were associated with significantly poorer TRM (Table). Stem cell source was not significant (p=0.157) in the model. Hemorrhage was reported as cause of death in 1/50 (2.0%) patients during P2 with PI-PC and 4/47 (8.5%) patients during P1 with C-PCs. Conclusions: Universal implementation of PI-PC in routine with extended storage to 7 days in P2 was associated with reduced TRM and better overall survival 100 days post HCT, despite transplantation of older patients with higher EBMT risk scores. Multivariate analysis revealed an adjusted hazard ratio of 0.433 (95% C.I. 0.262, 0.716) for TRM by 100 days, suggesting better outcomes in P2. This retrospective analysis at a single site indicated that PI-PC treated with amotosalen /UVA stored up to 7 days did not have a negative impact on TRM and OS in HCT recipients, and was an integral part of improving clinical outcomes at our institution. . Table. Disclosures Heim: Novartis: Research Funding. Irsch:Cerus Corporation: Employment, Equity Ownership. Lin:Cerus Corporation: Employment, Equity Ownership. Benjamin:Cerus Corporation: Employment, Equity Ownership. Corash:Cerus Corporation: Employment, Equity Ownership.

Cytogenetic Clonal Evolution in MDS Is Associated with Shifts towards Unfavorable Karyotypes According to IPSS and Shorter Overall Survival: A Study on 988 MDS Patients Studied Sequentially by Chromosome Banding Analysis

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 968-968 ◽  
Author(s):  
Claudia Haferlach ◽  
Melanie Zenger ◽  
Tamara Alpermann ◽  
Susanne Schnittger ◽  
Wolfgang Kern ◽  
...  
Keyword(s):  
Overall Survival ◽  
Chromosome Banding ◽  
Cox Regression ◽  
Clonal Evolution ◽  
Employment Equity ◽  
Cox Regression Analysis ◽  
Banding Analysis ◽  
Chromosome Banding Analysis ◽  
Equity Ownership ◽  
Acquired Abnormalities

Abstract Abstract 968 Background and Aim: The karyotype is one of the most important prognostic factors in MDS with respect to survival and evolution to AML and may change during the course of the disease. The aim of this study was to evaluate 1. the frequency of acquisition of additional chromosome abnormalities during the course of the disease (clonal evolution), 2. the pattern of acquired genetic abnormalities, 3. the association of karyotype at diagnosis and clonal evolution and 4. the impact of clonal evolution on transformation to AML and overall survival (OS). Patients and Methods: 988 MDS patients were evaluated by chromosome banding analysis (CBA) during the course of their disease. According to IPSS 729 (73.8%) cases showed a favorable karyotype, 146 (14.8%) patients an intermediate karyotype and 113 (11.4%) cases an unfavorable karyotype at first investigation. Progression to AML occurred in 180 of 988 patients during follow-up. Results: 2,454 chromosome banding analyses were performed in 988 cases (mean: 2.48 per case, range: 2–9). The median time between the first and the last evaluation was 12.5 months (range 1–60.6 months). Overall, in 171 of 988 patients (17.3%) clonal evolution was observed. Clonal evolution was detected between 1 and 56 months (median 14.3 months) after first evaluation and occurred later in patients with favorable than in patients with intermediate or unfavorable karyotype (mean 19.8 mo vs 15.5 mo vs 10.5 mo, favorable vs intermediate p=0.07, intermediate vs unfavorable p=0.05 and favorable vs unfavorable p<0.001). The abnormalities most frequently acquired during the course of the disease were +8, 7q−/−7, and gain of 21q detected in 29 cases each, followed by loss of 12p (n=22), 5q (n=14), 17p (n=19), and 20q (n=12). Other recurrently acquired abnormalities were +13 (n=12), +1q (n=12), +3q (n=12), −3q (n=10). Clonal evolution was strongly associated with cytogenetic IPSS category: Clonal evolution occurred in 100/729 cases with upfront favorable cytogenetics (13.7%), in 32/146 patients (21.9%) with upfront intermediate cytogenetics, but in 39/113 cases (34.5%) with upfront unfavorable cytogenetics (p<0.001). In 100 patients with favorable cytogenetics and clonal evolution karyotype was intermediate at second evaluation in 43 cases (43%), unfavorable in 25 cases (25%) and stayed favorable in the remaining 32 patients (32%). In 32 patients with intermediate cytogenetics and clonal evolution karyotype shifted to unfavorable at second evaluation in 11 cases (34.4%) and stayed intermediate in 21 patients (65.6%). Progression to AML was more frequent in patients with clonal evolution as compared to patients without (52/171 (30.4%) vs 128/817 (15.7%); p<0.001). In Cox regression analysis the IPSS karyotype at first evaluation, the IPSS karyotype at second evaluation, clonal evolution and progression to AML were associated with OS (relative risk: 2.12, 2.15, 1.87, and 6.6; p<0.001, p<0.001, p=0.011, p<0.001, respectively). In multivariate Cox regression analysis the IPSS karyotype at second evaluation and progression to AML were independently associated with shorter OS (relative risk: 2.0, and 6.1; p=0.013, p<0.001, respectively). Clonal evolution was associated with shorter OS (median 130.4 months vs not reached, OS at 5 years 72.3%vs 82.9%, p=0.01). Also in the subset of patients without transformation to AML outcome was inferior in patients with clonal evolution as compared to those without clonal evolution (OS at 5 years 78.2% vs 83.0%, p=0.05). Conclusions: 1. Clonal evolution was observed in 17.3% of patients with MDS. 2. The pattern of acquired abnormalities resembles the pattern observed in MDS at primary evaluation. 3. A higher frequency of clonal evolution and a shorter time to clonal evolution is observed in higher cytogenetic IPSS scores determined at first evaluation. 4. Clonal evolution is significantly associated with transformation to AML and shorter OS. 5. Sequential cytogenetic analyses allow the identification of subsets of MDS patients with a higher risk for transformation to AML and thus might guide treatment decisions in future. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Zenger:MLL Munich Leukemia Laboratory: Employment. Alpermann:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Modulation of the Clonal Composition in Relapsed CLL: A Study Based on Targeted Deep-Sequencing of ATM, BIRC3, NOTCH1, POT1, SF3B1, SAMHD1 and TP53

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1721-1721
Author(s):  
Sabine Jeromin ◽  
Wolfgang Kern ◽  
Richard Schabath ◽  
Tamara Alpermann ◽  
Niroshan Nadarajah ◽  
...  
Keyword(s):  
Deep Sequencing ◽  
Clonal Evolution ◽  
Gene Mutations ◽  
Wild Type ◽  
Mutation Load ◽  
Employment Equity ◽  
Time Points ◽  
Mutational Status ◽  
Equity Ownership ◽  
Time Point

Abstract Background: Relapse or refractory disease is a challenging clinical problem in the majority of chronic lymphocytic leukemia (CLL) patients. Treatment influences the clonal composition by selection and eventually induction of additional genetic abnormalities. Aim: To characterize the clonal evolution in relapsed CLL patients by deep-sequencing analysis of mutations in ATM, BIRC3, NOTCH1, POT1, SF3B1, SAMHD1 and TP53. Patients and Methods: Sequential samples of 20 relapsed CLL patients at three time-points were evaluated: A: at diagnosis (n=16) or in untreated state (n=4), B: at first relapse (n=20) and C: at second relapse (n=2). Patients were treated with diverse treatment schemes and had temporarily achieved either complete or partial remission during the course of the disease. The median time from diagnosis to first-line treatment was 13 months (1 - 169 months). All geneswere sequenced by a deep sequencing approach (Illumina, San Diego, CA). IGHV mutational status was determined by direct Sanger sequencing at time-point A. Chromosome banding analysis (CBA) and FISH data on del(17p), del(11q), trisomy 12 (+12), and del(13q) were available in 33/42 and 36/42 samples, respectively. Results: Initially, samples at first relapse were sequenced. Mutations in SF3B1 (6/20, 30%), TP53 (5/20, 25%), ATM (5/20, 25%), NOTCH1 (4/20, 20%), and SAMHD1 (3/20, 15%) were detected at high frequencies. No mutations were detected in BIRC3 and POT1. In total, 75% of cases presented with at least one mutation (Figure 1): 8 (40%) cases had one, 6 (30%) cases had two and one patient had three genes concomitantly mutated (mut). Patients were also analyzed for IGHV mutational status at diagnosis and presented with unmutated status at a frequency of 85% (17/20). Subsequently, samples from cases with mutations were analyzed at time-point A. In 12/15 (80%) cases the mutations at relapse were already detectable at time-point A with a similar load indicating presence of the main clone before and after chemotherapy. However, in 7/15 (47%) patients new gene mutations were acquired either additionally to existing mutations (n=4) or in previously wild-type cases (n=3). In 5/7 (71%) cases mutations were located in TP53. TP53 mut were the only mutations that were not detected in samples before treatment (sensitivity of 3%). Thus, TP53 mutations might have been initiated by chemotherapy or exist in a minor subclone subsequently selected by chemotherapy. Furthermore, only 4 cases had low-level mutations (3-6% mutation load) at diagnosis in either SAMHD1 or SF3B1 that eventually increased in their burden during disease course. Of note, in two patients a multibranching clonal evolution could be identified (#2, #9). For patient #2 three time-points were analyzed. At diagnosis 2 ATM mutations were detected with mutation loads of about 20%, each. In the course of the disease these mutations were lost, whereas SF3B1 mut showed a stable mutation load in all three time-points of about 40%. In contrast, mutation load of SAMHD1 increased over time from 4% to 87%. CBA was performed at diagnosis and detected independent clones with del(11q) and del(13q). Accordingly, del(11q) detected by FISH at diagnosis was lost and the percentage of cells with del(13q) increased from diagnosis to time-point C. Therefore, patient #2 shows different genetic subclones in parallel that were eradicated or selected by chemotherapy. In patient #9 two SF3B1 mutations were initially detected with the same mutation load of 10%. After treatment one mutation was lost, whereas the load of the second mutation increased indicating at least two different subclones with only one of them being sensitive to chemotherapy. This might be due to different additional aberrations. Indeed, CBA identified two clones: one with +12 alone and one in combination with del(13q). FISH revealed unchanged percentage of +12 at time-point B, whereas del(13q) positive cells were diminished. Conclusions: In 75% of relapsed CLL cases mutations in SF3B1, TP53, ATM, NOTCH1, and SAMHD1 are present at high frequencies. 80% of these mutations are already detectable before treatment initiation representing the main clone. Remarkably, TP53 mutations were the only mutations that were not detected before but only after chemotherapy. Figure 1. Distribution of gene mutations in 15 CLL cases with mutations at diagnosis or before treatment (D) and at relapse (R). Red = mutated, grey = wild-type, white = not analyzed. Figure 1. Distribution of gene mutations in 15 CLL cases with mutations at diagnosis or before treatment (D) and at relapse (R). Red = mutated, grey = wild-type, white = not analyzed. Disclosures Jeromin: MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Schabath:MLL Munich Leukemia Laboratory: Employment. Alpermann:MLL Munich Leukemia Laboratory: Employment. Nadarajah:MLL Munich Leukemia Laboratory: Employment. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

scholarly journals Characterization of Del(14)(q24q32) in Mature B-Cell Neoplasms By Array-CGH, Cytogenetics and Molecular Mutations

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3901-3901
Author(s):  
Sabine Jeromin ◽  
Wolfgang Kern ◽  
Torsten Haferlach ◽  
Susanne Schnittger ◽  
Claudia Haferlach
Keyword(s):  
B Cell ◽  
Cox Regression ◽  
Univariate Analysis ◽  
Comparative Genomic ◽  
Molecular Characteristics ◽  
Splenic Marginal Zone Lymphoma ◽  
Employment Equity ◽  
Cytogenetic Aberrations ◽  
Mutational Status ◽  
Equity Ownership

Abstract Background: Deletions of 14q occur recurrently in mature B-cell neoplasms at a low frequency of 1.5% (Reindl et al., BJH 2010). In about one-third of these cases breakpoints show a clustering at 14q24.1 (centromeric) and at 14q32.3 (telomeric). Limited genetic data is available on this rare subgroup. Aim: To characterize del(14)(q24q32) using array based comparative genomic hybridization (aCGH) and to analyze cytogenetical and molecular characteristics. Patients and Methods: 34 patients with mature B-cell neoplasms and del(14)(q24q32) by chromosome banding analysis were analyzed. Median age was 72 years (range: 45-94 years). Male:female ratio was 1.4:1. All cases were analyzed by aCGH (Agilent, Waldbronn, Germany) and for mutations in MYD88, NOTCH1, TP53, and SF3B1 as well as for IGHV mutational status by direct Sanger sequencing. IGHV mutated (mut) cases without stereotypic VH3-21 were classified as IGHV favourable (IGHV fav). For statistical analysis of CLL patients data were compared with a cohort of 1,136 untreated CLL patients without del(14q). Results: Patients with del(14)(q24q32) were immunophenotypically classified as follows: 26 (59%) had CLL (n=20) or CLL/PL (n=6), 6 (18%) had splenic marginal zone lymphoma (SMZL), one patient had two B-cell neoplasms (SMZL and CLL/PL: 66% and 10% infiltration, respectively) and one patient had monoclonal B-cell lymphocytosis that progressed to CLL within two years. Analysis with aCGH showed that centromeric breakpoints were detected within a region of 970 kb (69,135,775 - 70,106,558) and telomeric breakpoints localized within a region of 712 kb (105,618,017 - 106,329,974). The median length of the deletions was 36.9 Mb (range: 35.5-37.1). Interestingly, the same breakpoints were present in 4 patients, each: 69,248,772 - 106,329,974 (cluster 1) and 69,271,436 - 106,329,974 (cluster 2). Genes located at the centromeric breakpoint that may be activated by juxtaposition to the IGH locus include FUT8. Its expression contributes to cancer malignancy. Interesting candidate genes located within the deleted region are PPP1R13B (p53 interaction) and NUMB (NOTCH1 interaction). In addition to del(14)(q24q32) 39 cytogenetic aberrations were detected in 23 patients. Two changes were recurrent: trisomy 12 (+12; n = 14) and del(13q) (n = 3). All patients of cluster 1 had +12 (cluster 1 vs. non-cluster 1: 100% vs. 33%, p=0.022). Mutations were detected in TP53 (n = 5, 15%) and NOTCH1 (n = 12, 35%). In SF3B1 a variant and in MYD88 no mutation were found. All mutated cases were CLL or CLL/PL cases only (n.s.). No molecular or cytogenetic differences were detected between CLL or CLL/PL and SMZL. Additionally, the IGHV mutational status was determined in CLL and CLL/PL cases (unmutated in 18 (69%), mutated in 8 (31%)). Further statistical analyses were performed in cases with CLL or CLL/PL with vs. without del(14)(q24q32). NOTCH1 mut (42% vs. 12%, p<0,001) and +12 (46% vs. 14%, p<0.001) were more frequent in patients with del(14q) vs. without. Of note, NOTCH1 mut were not associated with +12 in patients with del(14q) (with vs. without +12: 21% vs. 45%, n.s.), whereas patients without del(14q) showed a significant association (31% vs. 9%, p<0.001). Furthermore, del(13q) was rare in patients with del(14q) (4% vs. 61%, p<0.001) and IGHV fav was detected in less cases (35% vs. 60%, p=0.014). In 978 cases (events = 373; del(14q): n=13, events = 11) data on time to treatment (TTT) was available. TTT was shorter in patients with vs. without del(14q) (4 vs. 95 months, p<0.001). This was also the case when TP53 disrupted (del(17p) and TP53 mut) and +12 cases were excluded from the del(14q) cohort (17 vs. 95, p<0.001). For Cox regression analyses cytogenetic aberrations were hierarchically classified as follows: del(14q), del(17p), del(11q), +12, del(13q) sole. Univariate analysis of cytogenetic subgroups and molecular mutations identified that del(14q), TP53 disrupted, del(11q), +12, NOTCH1 mut, and SF3B1 mut have significant negative impact on TTT and del(13q) sole and IGHV fav are positive prognosticators. Multivariate analysis showed independent impact of del(14q) (HR: 5.2, p<0.001), del(11q) (HR:1.5, p=0.037), SF3B1 mut (HR: 1.5, p=0.008), and IGHV fav (HR: 0.3, p<0.001). Conclusions: del(14)(q24q32) are deletions with a very low variability in breakpoints in mature B-cell neoplasms. In CLL patients they are associated with +12, NOTCH1 mut and independently with a very short TTT. Disclosures Jeromin: MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Systematic Analysis of Cell-of-Origin, Sequencing and Genomic Imbalances Identifies a Distinct Subset of Rituximab Treated Diffuse Large B-Cell Lymphoma with an Inferior Survival

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3910-3910
Author(s):  
Rajan Dewar ◽  
Julia Friedman ◽  
Daniel Xia ◽  
Timothy George Lens ◽  
Asha Guttapalli ◽  
...  
Keyword(s):  
Copy Number ◽  
Cell Lymphoma ◽  
Univariate Analysis ◽  
Gene Mutations ◽  
B Cell Lymphoma ◽  
Cell Of Origin ◽  
Employment Equity ◽  
Systematic Analysis ◽  
Genomic Imbalances ◽  
Equity Ownership

Abstract Introduction: Diffuse Large B-cell Lymphoma (DLBCL), exhibits a wide range of clinical and biological heterogeneity. While initially responsive to rituximab based therapy, more than 70% of patients relapse within 5 years. Two distinct subtypes of DLBCLs were previously identified by gene expression profiling, that correlated to the cell of origin (COO); currently an immunohistochemistry (IHC) surrogate (Hans algorithm) is used to identify this COO. However, the value of COO identification by IHC is being questioned in patients treated with rituximab based regimen. As such there is a need for the identification of additional prognostication markers. The purpose of the present study is to systematically characterize a cohort of patients with DLBCL using a variety of methods including COO by IHC, copy number changes by array-based comparative genomic hybridization, and single gene mutations by next generation sequencing. The goal is to identify prognostic molecular biomarkers that predict survival better than current methods such as COO in DLBCL patients treated with rituximab. Methods: Patients diagnosed between 2003 and 2011 and treated with R-CHOP/R-EPOCH at BIDMC were identified. IPI/R-IPI, ECOG performance status, overall survival data were collected from retrospective chart review. Molecular and immunohistochemical studies were performed on formalin-fixed paraffin embedded tissue (FFPE), which was obtained prior to initiation of therapy (at the time of diagnosis). aCGH: Using a previously developed assay for aCGH to detect genomic gain/loss from archival FFPE, we characterized each DLBCL sample for the presence or absence of 50 copy number variations (CNVs) from 32 common regions of overlapping genomic imbalances comprising 36 minimal common regions. The calling criteria were based on GISTIC defined peaks based on copy number data from three publicly available datasets: IS-172, IS-51HR, EMEXP-3463. Gene panels: Next Generation Sequencing (NGS) was performed using a targeted hybrid capture panel and run on a Miseq (Illumina, Inc.). Gene selection for the panel was based on frequently mutated genes reported in DLBCL, Follicular lymphoma (FL), and Mantle Cell Lymphoma (MCL). Also selected for the panel were genes involved in known dysregulated pathways, therapeutic targets, and genes mapped to sites of genomic gains or losses. Cell of Origin: Immunostains were performed for BCL6, CD10, MUM1, FOXP1, GCET and LMO2. The immunostains were blindly scored by three different pathologists. For this study, GCB vs ABC determination was made using the Hans algorithm (CD10, BCL6 and MUM1 expression). Statistics: Kaplan-Meier (KM) survival analysis was performed using R (version 3.2.1). Results: Data on an initial subset of 49 patients with comprehensive molecular, COO and clinical information is presented (and an additional ~100 case analysis is in progress). Average age of patients at the time of diagnosis was 63, with 23 males and 26 females. By univariate analysis, the COO was not significantly associated with survival in the patients treated with rituximab based regimen (KM p=0.678). By contrast, three of the CNVs were associated with survival by Kaplan Meier analysis (Table 1). Also four, gene mutations were significantly associated with poor survival by univariate analysis (Table 1). Table 1.Markers significantly associated with survival in rituximab treated patients with DLBCL. Ab28 predicted for better survival, whereas the presence of the other mutations or CNVs predicted for poor survival.Gene / CRp value (KM)Ab29 (D1p13.1)0.00787PIK3C2G0.00126PIM10.00499CD79B0.00567DTX10.0102Ab3(A1q21.1-q25.1(3))0.0193Ab28 (D1p36.32-p36.31)0.0211 Conclusions: Our systematic analysis of DLBCLs using multiple immunohistochemical and molecular methods identified mutational and copy number biomarkers predictive of survival in DLBCL patients, treated with rituximab in univariate analysis. Multivariate analysis and data from additional cases will reveal whether these molecular biomarkers can better predict patient survival, compared to current methods (such as COO by IHC). Disclosures Friedman: Cancer Genetics Inc.,: Employment, Equity Ownership. Guttapalli:Cancer Genetics, Inc.: Employment, Equity Ownership. Thodima:Cancer Genetics, Inc.: Employment, Equity Ownership. Kamalakaran:Cancer Genetics, Inc.: Employment, Equity Ownership. Houldsworth:Cancer Genetics Inc.,: Employment, Equity Ownership.

scholarly journals Impact of Somatic Gene Mutations on Response to Lenalidomide (LEN) in IPSS Lower-Risk Myelodysplastic Syndromes (MDS) Patients (Pts) without Del(5q) and Ineligible for or Refractory to Erythropoiesis-Stimulating Agents (ESAs)

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 225-225 ◽  
Author(s):  
Valeria Santini ◽  
Pierre Fenaux ◽  
Aristoteles Giagounidis ◽  
Uwe Platzbecker ◽  
Alan F List ◽  
...  
Keyword(s):  
Lower Risk ◽  
Research Funding ◽  
Response Rate ◽  
Single Gene ◽  
Gene Mutations ◽  
Advisory Board ◽  
Employment Equity ◽  
Mutation Status ◽  
Somatic Gene ◽  
Equity Ownership

Abstract Background: Somatic gene mutations occur in the majority of MDS pts; specific mutations and high mutation frequency have prognostic relevance (Papaemmanuil et al. Blood. 2013;122:3616-27). Evaluation of somatic mutations may support the diagnosis of MDS and guide treatment (Tx) selection. The phase 3 randomized MDS-005 study compared LEN and placebo (PBO) Tx in red blood cell transfusion-dependent (RBC-TD) non-del(5q) lower-risk MDS pts ineligible for or refractory to ESAs. Deletions in chromosome 5q are associated with a high response rate to LEN in MDS pts; however, no mutations have been definitively associated with a predictable clinical response to LEN in non-del(5q) MDS. Aim:To investigate the relationship between somatic gene mutations detected by targeted next-generation sequencing (NGS) and response and overall survival (OS) in lower-risk non-del(5q) MDS pts treated with LEN in the MDS-005 study. Methods: Eligible pts were: RBC-TD (≥ 2 units packed RBCs/28 days 112 days immediately prior to randomization) with International Prognostic Scoring System defined Low-/Intermediate-1-risk non-del(5q) MDS; ineligible for ESA Tx (serum erythropoietin > 500 mU/mL); or unresponsive or refractory to ESAs (RBC-TD despite ESA Tx with adequate dose and duration). 239 pts were randomized 2:1 to oral LEN 10 mg once daily (5 mg for pts with creatinine clearance 40-60 mL/min) or PBO. DNA was isolated from bone marrow mononuclear cells or whole blood collected at screening from a subset of pts who gave informed consent for this exploratory biomarker analysis and had adequate tissue for analysis. Targeted NGS of 56 genes was performed at Munich Leukemia Laboratory; average sequencing coverage was 2,000-5,000-foldand the variant allele frequency detection cutoff was 3%. Target regions varied by gene, including all exons to hotspots. For association tests, mutant variants (heterozygous or homozygous) were scored as 1 (mutant) or 0 (wildtype) for gene-level analyses. A Fisher exact test was used to test association of mutation status with response. Median OS was calculated by the Kaplan-Meier method. Hazard ratios and 95% confidence intervals were determined by a non-stratified Cox proportional hazards model. A log-rank test was used to test treatment effect with OS for single gene mutation status. Results: The biomarker cohort included 198 of 239 pts (83%; LEN n = 130, PBO n = 68). At least 1 mutation was detected in 30/56 (54%) genes and 173/198 (87%) pts. The most frequently mutated genes were SF3B1 (59%), TET2 (33%), ASXL1 (23%), and DNMT3A (14%); the most frequent co-mutations were SF3B1/TET2 (23%), SF3B1/DNMT3A (10%), SF3B1/ASXL1 (10%), and TET2/ASXL1 (9%) (Figure). Of 116 pts with SF3B1 mutations, 115 (99%) had ≥ 5% ring sideroblasts. The 56-day RBC transfusion-independence (RBC-TI) response rate was significantly lower in LEN-treated ASXL1 mutant pts vs wildtype pts (10% vs 32%, respectively; P = 0.031). At 168 days, the RBC-TI response rate was still lower in LEN-treated ASXL1 mutant pts vs wildtype pts (7% vs 22%); however, the difference was not significant (P = 0.101). LEN-treated DNMT3A mutant pts had a higher 56-day RBC-TI response rate vs wildtype pts (44% vs 25%); however, this difference did not reach significance (P = 0.133) due to the small sample size. RBC-TI response rate with LEN was similar regardless of total number of mutations per pt. Higher numbers of mutations were significantly associated (P = 0.0005) with worse median OS. Mutation in any of the genes associated with a negative prognosis reported by Bejar et al. (N Engl J Med. 2011;346:2496-506) was also significantly associated (P = 0.0003) with worse median OS.However, OS was not significantly different in LEN- vs PBO-treated pts based on any single gene mutation status. Conclusions: In this group of lower-risk RBC-TD non-del(5q) MDS pts, somatic mutations in genes recurrently mutated in myeloid cancers were detected in 87% of pts. SF3B1 mutations (alone or in combination) were most frequent and not associated with response to LEN. ASXL1 mutant pts had a significantly lower LEN response rate vs wildtype pts, whereas DNMT3A mutant pts had a trend for improved LEN response. Median OS was influenced by mutations, but not significantly modified by LEN. Determining predictive clinical markers for Tx response in non-del(5q) MDS pts remains challenging; nevertheless, there is a significant need to identify pt subsets who may be responsive to LEN Tx. Figure. Figure. Disclosures Santini: Novartis: Consultancy, Honoraria; Amgen: Other: advisory board; Onconova: Other: advisory board; Celgene: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria; Astex: Other: advisory board. Fenaux:Celgene, Janssen, Novartis, Astex, Teva: Research Funding; Celgene, Novartis, Teva: Honoraria. Giagounidis:Celgene Corporation: Consultancy. Platzbecker:Janssen-Cilag: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Celgene Corporation: Honoraria, Research Funding; Amgen: Honoraria, Research Funding; TEVA Pharmaceutical Industries: Honoraria, Research Funding. Zhong:Celgene Corporation: Employment, Equity Ownership. Wu:Celgene Corporation: Employment, Equity Ownership. Mavrommatis:Discitis DX: Membership on an entity's Board of Directors or advisory committees; Celgene Corporation: Employment, Equity Ownership. Beach:Celgene Corporation: Employment, Equity Ownership. Hoenekopp:Celgene Corporation: Employment, Equity Ownership. MacBeth:Celgene Corporation: Employment, Equity Ownership, Patents & Royalties, Research Funding.

Translocations Involving the IGH@locus Occur In 3.7% of Chronic Lymphocytic Leukemia and Are Associated with Unmutated IGHV Status and a Shorter Time to Treatment: A Study on 2,135 Cases

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3596-3596
Author(s):  
Claudia Haferlach ◽  
Frank Dicker ◽  
Susanne Schnittger ◽  
Wolfgang Kern ◽  
Torsten Haferlach
Keyword(s):  
Multivariate Analysis ◽  
Regression Analysis ◽  
Chronic Lymphocytic Leukemia ◽  
Cox Regression ◽  
Lymphocytic Leukemia ◽  
Employment Equity ◽  
Mutation Status ◽  
Cox Regression Analysis ◽  
Igh Locus ◽  
Equity Ownership

Abstract Abstract 3596 Introduction: Chronic lymphocytic leukemia (CLL) is a heterogeneous disease with a variable clinical course and a large spectrum of treatment options. Based on FISH data, a prognostic classification system has been established with 13q deletions as sole abnormality associated with a favorable prognosis and 17p and 11q deletions correlating with an unfavorable outcome. Recently, the combined evaluation of FISH data, IGHV mutation status and chromosome banding analysis (CBA) revealed that the impact of distinct genetic parameters differs with respect to overall survival (OS) and time to treatment (TTT). Thus far only few data is available on less frequent genetic abnormalities such as 14q deletions and translocations involving the IGH@ locus (tIGH). Therefore, we analyzed CLL with tIGH in detail with respect to frequency, partner genes and impact on prognosis. Methods/Patients: 78 CLL cases with tIGH were identified from 2,135 CLL sent to our laboratory for diagnostic work-up. All cases had been evaluated by immunphentotyping, FISH and CBA. Result: The most frequent tIGH was t(14;19)(q32;q13) (BCL3, n=21) followed by t(14;18)(q32;q21) (BCL2, n=19), t(8;14)(q24;q32) (CMYC, n=7) and t(11;14)(q13;q32) (CCND1, n=6). In the remaining 25 cases 5 recurrent translocations (t(2;14)(p13;q32), n=3; t(4;14)(p16;q32), FGFR3, n=2; t(11;14)(p15;q32), n=2; t(14;17)(q32;q25), n=2; and t(7;14)(q21;q32), n=2) were observed while the remaining 14 translocations were identified in single cases only. In 9/78 cases (11.5%) the tIGH was the sole abnormality. Recurrent additional chromosome abnormalities were +12 (n=7), del(13q) (n=9), del(11q) (n=3). A 17p deletion was observed in 1 case. In two cases tIGH was present only in a subclone and was a secondary abnormality occurring in addition to an del(11q) and a +12, respectively. CLL with tIGH were compared to 401 CLL without tIGH comprising all other genetic subgroups (subdivided according to Döhner et al.: del(17p) n=26, del(11q) n=42, +12 n=42, “normal” n=88, del(13q) sole n=177 and del(14q) n=26). An unmutated IGHV status was more frequent in CLL with tIGH as compared to all others (26/46 (54.3%) vs 128/353 (36.3%); p=0.023). For 53 cases with tIGH and all cases of the non-tIGH cohort clinical follow-up data was available. Median OS was 143.8 months (mo) in CLL with tIGH and 72.9 mo in patients with del(17p) while it was not reached in all other subgroups. In Cox regression analysis only del(17p) and mutated IGHV status were significantly associated with OS (p<0.0001, relative risk (RR)=7.0; p=0.014, RR=0.38). Median TTT was as follows: total cohort: 60.9 mo; tIGH: 27.8 mo; del(17p): 58.9 mo; del(11q): 19.7 mo; +12: n.r.; “normal” 63.9 mo; del(13q) sole: 83.0 mo and del(14q): 21.0 mo. In univariate Cox regression analysis the following parameters were significantly associated with shorter TTT: tIGH (p=0.004, RR=1.82), del(11q) (p<0.0001, RR=2.55), and del(14q) (p=0.007, RR=2.1), while del(13q) sole and mutated IGHV status were associated with longer TTT (p<0.0001, RR=0.40; p<0.0001, RR=0.23). In multivariate analysis including tIGH, del(11q), del(14q) and del(13q) sole all parameters retained their impact on TTT. However, if IGHV mutation status was included in the model only the mutated IGHV mutation status retained an impact on TTT (p<0.0001, RR=0.26). Next, patients with tIGH were subdivided according to their partner genes. Median OS was not reached in all subgroups, while median TTT was as follows: t(11;14): 101.2 mo, t(14;18): 47.9 mo, t(14;19): 11.0 mo, t(8;14): 18.5 mo and other partner genes: 27.8 mo. In univariate Cox regression analysis only t(14;19) was significantly associated with shorter TTT (p<0.001, RR=3.1). Including t(14;19) into multivariate analysis revealed a significant impact of both mutated IGHV mutation status and t(14;19) on TTT (p<0.0001, RR=0.286; p=0.004, RR=3.60). Conclusion: Translocations involving the IGH@ locus occur at low frequency in CLL. They are associated with unmutated IGHV status and a shorter TTT. TTT is especially short in cases with t(14;19). The prognostic impact of t(14;19) is independent of IGHV mutation status. In contrast CLL with t(11;14) and t(14;18) are neither associated with shorter OS nor shorter TTT. This data supports the application of CBA in CLL in order to identify all clinically relevant chromosomal aberrations, including those not detected by routine FISH analysis. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Dicker:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

TP53 Alterations In CLL - Parameters Influencing The Prognostic Impact: A Study On 3,988 Patients

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 865-865
Author(s):  
Claudia Haferlach ◽  
Frank Dicker ◽  
Sabine Jeromin ◽  
Sandra Weissmann ◽  
Andreas Roller ◽  
...  
Keyword(s):  
Cox Regression ◽  
Small Subset ◽  
Mutation Load ◽  
Employment Equity ◽  
Mutation Status ◽  
Cox Regression Analysis ◽  
Tp53 Mutations ◽  
Entire Cohort ◽  
Impact On Survival ◽  
Equity Ownership

Abstract Background In CLL, the TP53 gene may be inactivated by deletion and/or mutations. Most cases with 17p deletion also carry TP53 mutations on the second allele. However, in a subset of cases only one allele seems to be disrupted by either mutation or deletion. It is still a matter of debate whether monoallelic TP53 abnormalities have the same poor prognostic effect as biallelic alterations. Further, a small subset of patients with TP53 deletions harboring mutated IGHV genes were described to exhibit a slowly progressive disease without treatment indication for years. Aims In this study, we addressed the following questions: 1. Frequency of TP53 alterations: mutation and deletion. 2. Characterization of the TP53 altered subsets with respect to IGHV mutation status, other molecular mutations and cytogenetics. 3. Impact on survival. Patients and Methods 3,988 CLL patients were analyzed by DNA sequencing for TP53 mutations and by FISH for TP53 deletion status as well as for del(13q), del(11q) and +12. IGHV mutation status was determined in 3,505 patients. Further, SF3B1 (n=1,245), MYD88 (n=1,026), XPO1 (n=1,025), NOTCH1 (n=973), and FBXW7 (n=962) were analyzed by DNA sequencing. Results 488/3,988 (12.2%) harbored a TP53 mutation (TP53mut) and 308/3,988 (7.7%) patients showed a TP53 deletion (TP53del) by FISH. 268 cases (6.7%) showed both a TP53del and a TP53mut, while 220 cases (5.5%) harbored a TP53mut only and 40 (1.0%) a TP53del only. 20.5% of TP53mut cases harbored more than one TP53mut. The frequency of TP53mut and TP53del increased significantly with age (≤40 yrs: 2.4%/2.4%; 41-50 yrs: 7.5%/4.0%; 51-60 yrs: 12.4%/6.8%; 61-70 yrs: 12.1%/8.1%; 71-80 yrs: 13.4%/9.1%; >80 yrs: 16.0%/9.9%; p=0.006 and p=0.013, respectively). In the entire cohort, 1,428/3,505 (40.7%) cases showed an unmutated and 2,077/3,505 (59.3%) a mutated IGHV status. The lowest frequency of IGHV unmutated was observed in cases without TP53 alteration (1,148/3,094; 37.1%) and the highest in patients with both TP53mut and TP53del (156/201; 77.6%). The frequency was in between in patients with TP53mut sole (106/176; 60.2%) and TP53del sole (18/34; 52.9%). Patients with both TP53mut and TP53del as well as patients with TP53del sole had a significantly shorter overall survival (OS) compared to patients with TP53mut sole or patients without TP53 alteration (OS at 5 yrs: 40.2% vs. 36.4% vs. 68.8% vs 85.4%; p<0.001; TP53mut sole vs TP53wt: p=0.003). Next, we evaluated the impact of the TP53 mutation load on survival. Therefore, we divided patients into 10 subgroups according to their mutation load (increments of 10%). The OS of patients with a mutation load <20% (n=150) did not differ from patients with TP53wt, while a mutation load ≥20% was significantly associated with shorter OS (HR: 4.9, p<0.001). An unmutated IGHV status was associated with shorter OS in the total cohort (HR: 2.3, p<0.001). In the subset of patients with TP53wt an unmutated IGHV status was also an adverse prognostic factor (OS at 5 yrs: IGHV unmutated vs mutated: 80.3% vs 88.6%, p=0.007). This was true also in cases with TP53del sole (median OS: 12 months vs not reached, p=0.001). In contrast, in patients with either TP53mut sole or both TP53mut and TP53del the IGHV status had no impact on OS. In the entire cohort univariate Cox regression analysis revealed the following parameters to be significantly associated with OS: TP53mut (HR: 4.0), TP53mut ≥20% (HR: 4.9), TP53del (HR: 7.1), IGHV unmutated (HR: 2.3), age >60 yrs (HR: 3.3), del(11q) (HR: 2.1), del(13q) sole (HR: 0.6), SF3B1mut (HR: 2.5) (for all p<0.001), and NOTCH1mut (HR: 1.6, p=0.025). Multivariate Cox regression analysis including parameters significantly associated with OS in univariate analyses revealed the following factors to be independently associated with shorter OS: TP53del (HR: 4.2, p<0.001), TP53mut ≥20% (HR: 2.4, p=0.008), age >60 yrs (HR: 2.6, p<0.001), SF3B1mut (HR: 2.4, p<0.001), and del(11q) (HR: 2.2, p=0.002). Conclusions 1. TP53 alterations were observed in 13.2% of CLL patients, 6.7% showed both a deletion and a mutation, while 1% showed a deletion only and 5.5% a mutation only. 2. Both TP53 mutations and TP53 deletions are associated with an unmutated IGHV status. 3. TP53 deletions had the most adverse impact on survival, TP53 mutations had a significant impact on OS only if the mutation load was ≥20%. A small subset of patients with TP53 deletion sole and a mutated IGHV status seems to have a favorable outcome. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Dicker:MLL Munich Leukemia Laboratory: Employment. Jeromin:MLL Munich Leukemia Laboratory: Employment. Weissmann:MLL Munich Leukemia Laboratory: Employment. Roller:MLL Munich Leukemia Laboratory: Employment. Worseg:MLL Munich Leukemia Laboratory: Employment. Alpermann:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kohlmann:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Comprehensive Analysis Of U2AF1 Mutations In 843 Patients With Myeloid Neoplasms With Respect To Other Genetic Alterations

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4973-4973
Author(s):  
Manja Meggendorfer ◽  
Christiane Eder ◽  
Sabine Jeromin ◽  
Claudia Haferlach ◽  
Wolfgang Kern ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Hot Spots ◽  
Melting Curve ◽  
Gene Mutations ◽  
Next Generation ◽  
Employment Equity ◽  
Total Cohort ◽  
Equity Ownership ◽  
Generation Sequencing ◽  
Splicing Machinery

Abstract Introduction Genes affecting the splicing machinery have been found to be frequently mutated in MDS patients. U2AF1 codes for one of these splicing components, showing two distinct mutational hot spots at amino acids Ser34 and Gln157. Mutations in U2AF1 induce global abnormalities in RNA splicing, producing intron containing unspliced RNAs. U2AF1 has been shown to be most frequently mutated in MDS cases (7-11%), but was so far investigated only in small subsets of AML and MPN and was found rarely mutated. Aim To determine the frequency of U2AF1 mutations (U2AF1mut) in different myeloid entities and to evaluate the correlation of U2AF1mut with other gene mutations, cytogenetics and clinical features. Patients and Methods The total cohort consisted of 843 patients, whereof 74 were diagnosed as AML, 201 as MDS, 243 as MPN, and 325 as MDS/MPN overlap. 331 patients were female, 512 male. Cytogenetics was available in 830 patients and these were grouped by the following karyotypes: normal karyotype (n=561), +8 (n=39), -7 (n=15), del(20q) (n=95), -Y (n=29), other aberrations (n=59), and complex karyotype (n=32). Based on the previously described association of U2AF1mut with del(20q) there was an intended selection bias to this abnormality. Mutational analyses for U2AF1 were performed by either melting curve analyses or next generation sequencing. In subcohorts we investigated mutations in ASXL1 (n=505), CBL (n=647), CEBPA (n=68), CSF3R (n=213), DNMT3A (n=260), ETV6 (n=129), EZH2 (n=355), FLT3-ITD (n=352), FLT3-TKD (n=239), IDH1/2 (n=367 and 286, respectively), JAK2 (n=681), KITD816 (n=244), KRAS (n=393), MLL-PTD (n=384), MPLW515 (n=612), NPM1 (n=477), NRAS (n=509), RUNX1 (n=516), SETBP1 (n=336), SF3B1 (n=839), SRSF2 (n=784), TET2 (n=428), and TP53 (n=239) by Sanger sequencing, next generation sequencing, gene scan, or melting curve analysis. Results In the total cohort we detected U2AF1 mutations in 55/843 (6.5%) patients, the two mutational hot spots were equally affected with 29 p.Ser34 and 26 p.Gln157 mutations, respectively. Mutation frequencies were 10.9% in MDS, 9.5% in AML, 7.1% in MDS/MPN overlap and 1.2% in MPN. U2AF1mut patients were older (median: 72.6 vs. 71.8 years; p=0.012), the mutation was more frequent in males (42/512 (8.2%) vs. 13/331 (3.9%) in females; p=0.015) and associated with lower hemoglobin levels (median: 9.5 vs. 11.0g/dL; p<0.001), and platelet counts (median: 78x109/L vs. 179x109/L; p=0.002). Regarding cytogenetics we found a high association of U2AF1mut to del(20q): in 18 of 95 cases (18.9%) with del(20q) a U2AF1 mutation was detected compared to 37 U2AF1mut in 735 cases (5.0%) with any other karyotype (p<0.001). This was true for AML (5/16 vs. 2/56; p=0.005), MDS (11/49 vs. 11/150; p=0.007) and MDS/MPN overlap cases (1/8 vs. 21/309; p=0.441). In contrast in MPN none of the 21 del(20q) patients showed a U2AF1 mutation compared to 18/74 in all other entities (p=0.01). Mutations in the two other genes of the splicing machinery, SF3B1 and SRSF2, occurred in 122/839 (14.5%) and 198/784 (25.3%) cases and were mutually exclusive with U2AF1mut. Only one case each showed an U2AF1mut and a SF3B1 (p=0.002) or SRSF2 (p<0.001) mutation. We furthermore analyzed a number of other gene mutations frequently mutated in myeloid entities and their association to U2AF1mut. There was no correlation to mutations in NPM1, FLT3-ITD and FLT3-TKD, MLL-PTD, and CEBPA in AML patients. In MDS patients there was also no correlation to mutations in ASXL1,ETV6, EZH2, TP53, RUNX1, NRAS, and KRAS. This was also true for JAK2, MPL, CBL, and TET2 mutations in MPN. However in MDS/MPN overlap patients U2AF1mut were more frequently found in cases with ASXL1mut (14/115 (12.2%) in ASXL1mut vs. 7/179 (3.9%) in ASXL1wt; p=0.01) and together with KITD816mut (3/10 (30%) in KITD816mut vs. 15/212 (7%) in KITD816wt; p=0.038). Conclusion 1) U2AF1 is most frequently mutated in MDS, followed by AML and MDS/MPN overlap and in contrast rarely mutated in MPN. 2) U2AF1mut highly correlates with del(20q) in MDS, AML and MDS/MPN overlap but not in MPN cases. 3) In MDS/MPN overlap U2AF1mut associates significantly with ASXL1mut and KITD816mut. Disclosures: Meggendorfer: MLL Munich Leukemia Laboratory: Employment. Eder:MLL Munich Leukemia Laboratory: Employment. Jeromin:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Impact of 1p Deletions in Myelodysplastic Syndromes and Secondary AML Arising from Myelodysplastic Syndromes

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2006-2006
Author(s):  
Claudia Haferlach ◽  
Anna Stengel ◽  
Manja Meggendorfer ◽  
Wolfgang Kern ◽  
Torsten Haferlach
Keyword(s):  
Myelodysplastic Syndromes ◽  
Negative Impact ◽  
Chromosome 1 ◽  
Published Data ◽  
Employment Equity ◽  
Secondary Aml ◽  
Micro Rnas ◽  
Genomic Array ◽  
1P Deletion ◽  
Equity Ownership

Background: Deletions in the short arm of chromosome 1 are rare, recurrent abnormalities in Myelodysplastic syndromes (MDS) and are observed as the sole abnormality in 0.2% (Schanz et al. JCO 2012). So far no comprehensive characterization of this subset has been performed. Aim: The aim of this study was to characterize MDS and secondary AML evolving from MDS harboring a 1p deletion with respect to 1) accompanying cytogenetic and molecular genetic abnormalities, 2) the size of the 1p deletion and the minimal deleted region. Patients and Methods: 50 cases with MDS (de novo MDS: n=38, t-MDS: n=8) and secondary AML evolving from MDS (n=4) harboring a 1p deletion were selected for analysis. All cases were evaluated by chromosome banding analysis. From 30 cases sufficient material was available to perform genomic array analysis (SurePrint G3 ISCA CGH+SNP Microarray, Agilent, Waldbronn, Germany) and amplicon sequencing to detect mutations in ASXL1, CBL, CSF3R, CSNK1A1, DNMT3A, ETNK1, ETV6, EZH2, GATA1, IDH1, IDH2, JAK, KIT, KRAS, MPL, NPM1, NRAS, RUNX1, SETBP1, SF3B1, SRSF2, TET2, TP53, and U2AF1. Variants of unknown significance were excluded from statistical analysis. Results: 62% were male and median age was 75 years (range: 35 - 91). The 1p deletion was the sole chromosomal abnormality in 5/50 cases (10%) and was accompanied by one, two and more than two additional aberrations in 12 (24%), 15 (30%), and 18 (36%) cases, respectively. In total 129 chromosome abnormalities were observed in addition to the 1p deletion (median per patient: 2, range: 0-9). Of these only 10 were balanced, while 119 were unbalanced abnormalities leading to gain or loss of chromosomal material. Loss of 1p was most frequently accompanied by del(5q) (n=24; 48%), +8 (n=20; 39%), 7q-/-7 (n=11; 22%), del(17p) (n=5; 10%), and -Y (n=3; 6%). In 15 cases (29%) a duplication of the short arm of chromosome 1 harboring the 1p deletion was observed. Genomic array analyses revealed a median size of the 1p deletion of 25 MB (range: 13-34 MB). A minimal deleted region encountered in all 30 evaluable patients ranged from genomic position 17,872,935 to 24,285,861 encompassing 72 genes (e.g. E2F2, ID3, PAX7, UBR4, ZBTB40) and 10 micro RNAs. One, 2, 3, and 4 mutations were present in 10 (33%), 8 (26%), 5 (17%) and 2 (7%) cases, respectively. No mutations in any of the analyzed genes were observed in 5 cases (17%). Mutations were detected in SF3B1 (16%), TET2 (16%), ASXL1 (13%), DNMT3A (13%), EZH2 (13%), NRAS (13%), SRSF2 (13%), TP53 (13%), JAK2 (10%), KRAS (7%), U2AF1 (7%), CALR (3%), IDH2 (3%), MPL (3%), RUNX1 (3%), and SETBP1 (3%). No mutations were detected in CBL, CSF3R, CSNK1A1, ETNK1, ETV6, GATA1, IDH1, KIT, and NPM1. Compared to published data (Bejar et al. NEJM 2012, Papaemmanuil et al. Blood 2013, Haferlach et al. Leukemia 2014) SF3B1 (20-30%) and TET2 (20-30%) mutations seem to be less frequent and mutations in TP53 (6-8%), EZH2 (5-7%), NRAS (4-5%) and KRAS (1-3%) more frequent in MDS with 1p deletion compared to an unselected MDS cohort. Patients harboring either a 1p deletion as the sole abnormality or a duplication of the deleted chromosome 1 (n=20) had an excellent prognosis (3 year overall survival (OS): 100%), while patients with a 1p deletion accompanied by -7/7q- (n=11) had a very poor outcome (3 year OS: 0%; p<0.001). Patients with 1p deletion and other accompanying cytogenetic abnormalities (n=19) had an intermediate outcome (3 year OS: 34%, p=0.03, p=0.003). Conclusions: 1) Interstitial deletions in the short arm of chromosome 1 are rare recurrent abnormalities in MDS. 2) 5q deletion, +8, -7/7q- are frequently observed in addition to 1p deletion. 3) Both MDS with a 1p deletion as the sole abnormality and MDS with a duplication of deleted chromosome 1 are associated with very favorable outcome. 4) Accompanying abnormalities, especially -7/7q- have a negative impact on outcome. Disclosures Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Stengel:MLL Munich Leukemia Laboratory: Employment. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

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