About 17% of AML with NPM1 mutations Show a Specific Pattern of Chromosome Aberrations but These Cases Do Not Differ Prognostically from AML with NPM1 Mutations Carrying a Normal Karyotype

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2527-2527
Author(s):  
Claudia Haferlach ◽  
Susanne Schnittger ◽  
Tamara Weiss ◽  
Wolfgang Kern ◽  
Brunangelo Falini ◽  
...  
Keyword(s):  
Chromosome Aberrations ◽  
Chromosome Banding ◽  
Who Classification ◽  
De Novo ◽  
Normal Karyotype ◽  
Prognostic Impact ◽  
Banding Analysis ◽  
Chromosome Banding Analysis ◽  
Additional Chromosome ◽  
De Novo Aml

Abstract AML with mutated nucleophosmin gene (AML NPM1mut) usually carries a normal karyotype and will be suggested as a provisional entity in the new WHO classification. Thus far, the impact of chromosome aberrations in AML NPM1mut has not been evaluated in detail. Aim of this study was to determine the incidence and prognostic impact of clonal chromosome aberrations in NPM1mut. We further compared this pattern to additional aberrations in AML with recurrent genetic aberrations: t(8;21)(q22;q22), inv(16)(p13q22)/t(16;16)(p13;q22), t(15;17)(q22;q12) and 11q23-abnormalities leading to an MLL-rearrangement. In total 415 AML (de novo AML: 392, s-AML: 11, t-AML: 12) showing an NPM1 mutation were analyzed by chromosome banding analysis. 71 of these showed clonal chromosome aberrations (17.1%; de novo AML: 63 (16.1%), s-AML: 5 (45.5%), t-AML: 3 (25%); de novo AML vs. s-AML: p=0.024). Overall, 111 chromosome aberrations were observed. The most frequent abnormalities were +8 (n=30), −Y (n=10), +4 (n=9), del(9q) (n=5), +21 (n=4), −7 (n=3), +5 (n=2), +10 (n=2), +13 (n=2),+18 (n=2), del(12p) (n=2), del(20q) (n=2), other non-recurrent balanced aberrations (n=6), other non-recurrent unbalanced aberrations (n=32). For comparison 63 AML with t(8;21), 37 cases with inv(16)/t(16;16), 83 patients with t(15;17) and 83 AML showing a 11q23/MLL-rearrangement were evaluated. 44 (69.7%), 13 (35.1%), 39 (47%), and 28 (43.1%) cases showed clonal chromosome aberrations in addition, respectively. Therefore, additional chromosomal aberrations are more frequent in all these subgroups than in the AML NPM1mut. Similar to NPM1mut cases +8 (n=2), −X/Y (n=32), +4 (n=2), and del(9q) (n=10) were observed. The only other recurrent additional aberrations was del(11q) (n=2). In inv(16)/t(16;16) we also found +8 (n=5) and −Y (n=1). The only other recurring additional aberrations were +22 (n=6) and del(7q) (n=2). In AML with t(15;17) recurring additional abnormalities were +8 (n=12), −Y (n=3), del(9q) (n=2), ider(17)(q10) t(15;17) (n=7). AML with 11q23/MLL-rearrangement showed +4 (n=2), +8 (n=8), +13 (n=2), +19 (n=4), +21(n=4), +22 (n=2), −Y (n=1). Thus, chromosome aberration in AML NPM1mut share many overlaps to those in AML with recurrent aberrations. Furthermore, the prognostic impact of chromosome aberrations in AML NPM1mut was evaluated. No difference with respect to overall survival (OS) and event-free survival (EFS) was observed between AML NPM1mut with a normal (n=344) and an aberrant karyotype (n=71) (OS at 2 yrs 78% vs. 81%, p=0.969; EFS at 2 yrs 40% vs. 50%, median EFS 544 days vs. 522 days, p=0.253). The FLT3-ITD status was available in 400 cases. 127 (38%) of 334 cases with a normal karyotype showed a FLT3-ITD, while in only 16 (24%) of 66 cases with an aberrant karyotype a FLT3-ITD was observed (p=0.035). While the negative prognostic impact of additional FLT3-ITD was confirmed in our cohort, the presence of chromosome aberrations did not influence prognosis neither in the FLT3-ITD negative nor in the FLT3-ITD positive subgroup. In addition, 31 patients with AML NPM1mut were analyzed by chromosome banding analysis at diagnosis and at relapse (median time diagnosis to relapse: 301 days (range: 71–986). 22 cases (71%) showed a normal karyotype both at diagnosis and relapse. In 4 cases a normal karyotype was observed at diagnosis and an aberrant karyotype at relapse (del(9q) (n=2), t(2;11) (n=1), inv(12) (n=1)). One case with +8 at diagnosis showed +8 also at relapse. One case with +4 at diagnosis showed +4 and additional aberrations at relapse. In 1 case clonal regression was observed (+21 -> normal). One case with an unbalanced 1;3-translocation at diagnosis showed a der(17;18) (q10;q10) at relapse and one case with −Y at diagnosis showed a del(3p) at relapse. In conclusion: 1. Frequency of additional chromosome aberrations is low in AML NPM1mut as compared to other genetically defined WHO entities. 2. The pattern of additional chromosome aberrations is overlapping between the 5 groups analyzed. 3. Chromosome aberrations observed at diagnosis in AML NPM1mut do not influence prognosis in comparison to AML NPM1mut with normal karyotype. 4. The karyotype is stable in most AML NPM1mut patients at diagnosis and at relapse. These results point to chromosomal aberrations occurring in AML NPM1mut as secondary events and further support inclusion of AML NPM1mut as a provisional entity in the new WHO classification.

The Prognostic Impact of High EVI1 expression In AML Is Due to Its Close Correlation to Rearrangements Involving EVI1 or MLL, which Are Cytogenetically Cryptic In a Subset of Patients: a Study on 332 Cases.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1676-1676
Author(s):  
Claudia Haferlach ◽  
Vera Grossmann ◽  
Melanie Zenger ◽  
Tamara Alpermann ◽  
Alexander Kohlmann ◽  
...  
Keyword(s):  
Chromosome Banding ◽  
Normal Karyotype ◽  
Fish Analysis ◽  
Prognostic Impact ◽  
Employment Equity ◽  
Npm1 Mutation ◽  
Banding Analysis ◽  
Chromosome Banding Analysis ◽  
Equity Ownership ◽  
The Impact

Abstract Abstract 1676 Introduction: High EVI1 expression has been proposed as a negative prognostic factor in AML. An association between high EVI1 expression and distinct cytogenetic subgroups, such as 3q26-rearrangements, MLL-rearrangements and -7/7q- have been reported. Both 3q26- and MLL-rearrangements can be difficult to detect by chromosome banding analyses or may even be cytogenetically cryptic in a subset of patients due to limited resolution. Therefore, only studies using FISH for the detection of cryptic EVI1- or MLL-rearrangements can clarify their frequencies in AML with elevated EVI1 expression. Methods/Patients:: The study cohort was composed of 332 AML cases with a) normal karyotype (NK) (n=211), b) -7/7q- (n=77), and for comparison c) 3q26-rearrangements (n=38), and d) MLL-rearrangement (n=6). In all cases EVI1 expression was investigated using quantitative PCR calculating a % EVI1/ABL1 expression. In all cases FISH for EVI1 rearrangement was performed in addition to chromosome banding analysis. Cases with high EVI1 expression were also analyzed for MLL rearrangements by FISH. Results: In the total cohort, EVI1 expression varied between 0 and 1614 (median: 21.1). The highest EVI1 expression was measured in cases with cytogenetically identified 3q26-rearrangements (range: 6.1–566.4; median: 81.9) and in AML with MLL-rearrangements (range: 46.7–831; median: 239). The EVI1 expression was significantly lower in AML with NK (range: 0–1614; median: 0.5, p<0.001) and AML with -7/7q- (range: 0.03–199; mean: 34.5; median: 10.7, p<0.001). In the subgroup of cases with NK 4 MLL-rearrangements (1.9%) were detected by FISH and subsequently verified by fusion gene specific PCR. In addition, 4 cases with cryptic EVI1-rearrangements (1.9%) were identified by FISH analysis. Further genetic analysis revealed that these were due to t(3;8)(q26;q24) (n=2) and t(3;21)(q26;q11) (n=1). In one case, the EVI1-rearrangement could not be further analyzed due to lack of material. In the -7/7q- cohort 14/77 cases (18.2%) with cytogenetically cryptic EVI1 rearrangement including 3 novel recurrent abnormalities were detected: t(3;21)(q26;q11) (n=3), inv(3)(p24q26) (n=4) and t(3;8)(q26;q24) (n=2). In 5 cases FISH analysis revealed that the 7q- was not caused by an interstitial deletion but due to an unbalanced rearrangement between chromosomes 7 and 3: der(7)t(3;7)(q26;q21). In these 5 cases high-resolution SNP microarray were performed and revealed breakpoints in the CDK6 gene and centromeric of the EVI1 gene. Further mutation screening revealed that none of the cases with EVI1- or MLL-rearrangement harboured mutations in NPM1 or CEPBA. In 254 cases clinical follow-up data was available. Different cut-off levels of EVI1 expression were tested, and a cut-off at 30% EVI1/ABL1 expression was the lowest level that had a significant impact on outcome. Separating the cohort at this cut-off into high EVI1 (n=67) and low EVI1 expressors (n=187) showed a shorter EFS in patients with high EVI1-expression (p=0.001; relative risk (RR)=1.87, median EFS 6.2 vs 15.0 months (mo)), while no impact on OS was observed. When the same analyses were performed with respect to EVI1-rearrangements we observed both a significantly shorter EFS in cases with EVI1-rearrangement (n=39) vs all others (n=215) (p=0.001; RR=2.03, median EFS 4.6 vs 15.0 mo) and a significantly shorter OS (p=0.026; RR=1.73, median OS 10.1 vs 26.3 mo). Analyzing the impact of high EVI1 expression separately in the cohort without EVI1 rearrangement revealed no impact of EVI1 expression on EFS. Conclusions: The negative prognostic impact of high EVI1 expression is strongly associated with EVI1- or MLL-rearrangements and is absent in AML without EVI1- and MLL-rearrangement. Applying FISH in addition to chromosome banding analysis we identified cryptic rearrangements in 3.8% of AML with normal karyotype and in 18.2% of AML with -7/7q-, including 3 novel recurrent cytogenetically cryptic EVI1-rearrangements. This data supports the routine performance of FISH screening for EVI1- and MLL-rearrangements in patients with normal karyotype or 7q-/-7 and without NPM1 mutation and CEPBA mutation to assign patients to the correct biologic entity. The postulated independent prognostic impact of EVI1 expression should be tested further including this laboratory workflow as these parameters may have important impact on prognosis and future treatment strategies. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Grossmann:MLL Munich Leukemia Laboratory: Employment. Zenger:MLL Munich Leukemia Laboratory: Employment. Kohlmann: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.

Complex Aberrant Karyotypes Are Associated with Dismal Prognosis in CLL: Chromosome Banding Analysis Provides Important Prognostic Information in CLL in Addition to Interphase-FISH

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3132-3132
Author(s):  
Claudia Haferlach ◽  
Frank Dicker ◽  
Tamara Weiss ◽  
Susanne Schnittger ◽  
Wolfgang Kern ◽  
...  
Keyword(s):  
Chromosome Banding ◽  
Normal Karyotype ◽  
Prior Treatment ◽  
Prognostic Impact ◽  
Prognostic Information ◽  
Mutation Status ◽  
Interphase Fish ◽  
Cytotoxic Treatment ◽  
Banding Analysis ◽  
Chromosome Banding Analysis

Abstract CLL is a heterogeneous disease with a variable clinical course. Today, therapeutic decisions are based on clinical stage, prognostic information obtained by FISH analyses on interphase nuclei and determination of the IgVH mutation status. However, additional information might be obtained from chromosome banding analysis (CBA) which provides more details on genetic aberrations. Thus far, in CLL data from CBA have been scarce due to the low proliferative activity in vitro. We improved the cultivation technique using an immunostimulatory CpG-oligonucleotide DSP30 and IL-2 leading to a high success rate of CBA in routine diagnostics. Clinical follow-up was available in 533 CLL patients investigated in parallel with CBA and interphase-FISH with probes for the detection of trisomy 12, IGH-rearrangements and deletions of 6q21, 11q22.3 (ATM), 13q14 (D13S25 and D13S319) and 17p13 (TP53). Diagnosis of CLL was established by standard criteria based on immunophenotyping. In 463/533 cases IgVH mutation status was also available. 298 cases were analyzed at diagnosis (cohort 1), 121 during the course of their disease without prior treatment (cohort 2), 85 patients had received cytotoxic treatment prior to analysis (cohort 3) and for 29 cases no data were available with respect to prior treatment. First, we focused on the subset of patients who showed no aberrations as determined by FISH (n=120) and defined based on CBA 2 groups: normal karyotype (n=80), aberrant karyotype (n=40). No significant differences were observed with respect to OS or time to treatment (TTT). We then focused on complex aberrant karyotypes (3 or more clonal abnormalities). These are rarely found based on FISH diagnostics: we detected 22 cases (4.1%) that showed 3 or more aberrations based on FISH only as compared to 109 cases (20.5%) based on CBA. In detail, a complex aberrant karyotype was observed with comparable frequencies in the two cohorts analyzed at diagnosis (56/295, 19%) and during the course of their disease without prior treatment (22/123, 17.9%), while it was significantly more often found in the cohort analyzed after cytotoxic treatment (31/86, 36.0%; p=0.002). In both cohorts analyzed prior to any treatment patients with a complex aberrant karyotype had a significant shorter overall survival (p=0.042, HR=2.7 and p=0.003, HR=6.1). As TP53 deletions are associated with a complex aberrant karyotype and are a strong negative prognostic factor per se we analyzed the prognostic impact of complex aberrant karyotype in relation to TP53 deletions. Therefore, CLL patients analyzed at diagnosis with a complex aberrant karyotype by CBA (n=56) were subdivided into cases with TP53 deletion (n=17) versus without TP53 deletion (n=39) in FISH. TTT did not differ significantly between complex aberrant cases with or without TP53 deletion but was significantly shorter for both groups as compared to cases with 13q deletion or normal karyotype (n=135) (p=0.05 and p=0.02). Next, cases of cohort 1 with loss of 13q14 were divided based on CBA into 3 subgroups: as the sole abnormality (n=91), plus one additional abnormality (n=24), and plus 2 or more additional abnormalities (i.e. complex) (n=32). Also for these entities TTT was significantly shorter for subgroups 2 and 3 as compared to subgroup 1 (p=0.022, HR=2.5; p=0.001, HR=1.8). In conclusion, CBA allows to identify patients within the good prognostic FISH group del(13q) sole, who show a shorter TTT if additional abnormalities are identified by CBA. Even more striking, CBA defines a new subgroup of CLL with complex aberrant karyotype which shows a shorter TTT independent of the TP53 deletion status as detectable by FISH.

Interphase FISH and Comparative Genomic Hybridization Performed in Addition to Chromosome Banding Analysis in AML with Normal Karyotype Detect Prognostically Relevant Chromosome Abnormalities.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2016-2016 ◽  
Author(s):  
Claudia Schoch ◽  
Mirjam Klaus ◽  
Susanne Schnittger ◽  
Wolfgang Hiddemann ◽  
Wolfgang Kern ◽  
...  
Keyword(s):  
Comparative Genomic Hybridization ◽  
Chromosome Banding ◽  
Normal Karyotype ◽  
Trisomy 13 ◽  
Comparative Genomic ◽  
Genomic Hybridization ◽  
Interphase Fish ◽  
Banding Analysis ◽  
Chromosome Banding Analysis

Abstract In AML karyotype abnormalities are not detected in 40 to 45% of cases using classical chromosome banding analysis. For several reasons false negative results might occur in chromosome banding analysis: 1. no proliferation of the aberrant clone in vitro, 2. low resolution due to technical problems or limitations of the method itself, 3. real cryptic rearrangements. In order to determine the proportion of “false negative” karyotypes by chromosome banding analysis we conducted a study using interphase-FISH and comparative genomic hybridization in addition to chromosome banding analysis. In total, chromosome banding analysis have been performed in 3849 AML at diagnosis. Of these 1748 showed a normal karyotype (45.4%). Out of these in 3 cases cytomorphology revealed an APL and in 2 cases an AML M4eo. Using interphase FISH with a PML-RARA or CBFB probe we detected cryptic PML-RARA or CBFB-rearrangements, respectively, in all 5 cases, which were cytogenetically invisible due to submicroscopic insertions. 480 cases of AML with normal karyotype were analyzed for MLL gene rearrangements using FISH with an MLL-probe. 11 cases with a cryptic MLL-rearrangement were detected (FAB-subtypes: M5a: 7, M2: 2, M0: 2). In 273 patients interphase-FISH screening with probes for ETO, ABL, ETV6, RB, P53, AML1 and BCR was performed. In 6 out of 273 (2.2%) pts an abnormality was detectable. In two cases the aberrant clone did not proliferate in vitro: 1 case each with monosomy and trisomy 13. Due to limitations of resolution in chromosome banding analysis translocations or deletions of very small chromosome fragments were only detected with FISH in n=4 cases (ETV6 rearrangements: t(11;12)(q24;p13), t(12;22)(p13;q12), ETV6 deletions: del(12)(p13), n=2). Like interphase-FISH comparative genomic hybridization (CGH) does not rely on proliferating tumor cells but in contrast to interphase-FISH allows the detection of all genomic imbalances and not only of selected genomic regions. Therefore, we selected 48 cases with normal karyotype and low in vitro proliferation (less than 15 analyzable metaphases in chromosome banding analysis). In 8 of 48 cases (16.7%) an aberrant CGH-pattern was identified which was verified using interphase-FISH with suitable probes. In 3 cases a typical pattern of chromosomal gains and losses observed in complex aberrant karyotypes was detected. In one case each a trisomy 4 and 13 was observed, respectively. In one case trisomy 13 was accompanied by gain of material of the long arm of chromosome 11 (11q11 to 11q23). One case each showed loss of chromosome 19 and gain of the long arm of chromosome 10, respectively. In conclusion, CGH in combination with interphase-FISH using probes for the detection of balanced rearrangements is a powerful technique for identifying prognostically relevant karyotype abnormalities in AML assigned to normal karyotype by chromosome banding analysis. Especially this is true in cases with a low yield of metaphases and in AML with a high probability of carrying a specific, cytogenetically cryptic fusion-gene. Thus, in these cases interphase-FISH and CGH should be performed in a diagnostic setting to classify and stratify patients best.

AML with Recurrent Genetic Abnormalities and AML with Myelodysplasia-Related Changes Account for Three-Quarters of Previously Untreated Pediatric AML-the Results of a Nation-Wide Central Review in Japan-

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4899-4899
Author(s):  
Akitoshi Kinoshita ◽  
Hayato Miyachi ◽  
Hiromichi Matsushita ◽  
Tomohiko Taki ◽  
Miharu Yabe ◽  
...  
Keyword(s):  
Who Classification ◽  
De Novo ◽  
Conflicts Of Interest ◽  
Prognostic Significance ◽  
Prognostic Impact ◽  
Genetic Abnormalities ◽  
Precise Diagnosis ◽  
De Novo Aml ◽  
Pediatric Aml ◽  
Mixed Phenotype

Abstract Abstract 4899 [Background] The WHO classification has been widely accepted among physicians who are engaged in treating pediatric AML patients. In 2008, the revised WHO classification has expanded the two categories in AML; AML with recurrent genetic abnormalities and AML with myelodysplasia-related changes. The epidemiology and prognostic significance of these refined categories remains to be explored in children. [Methods] JPLSG AML-05 is a nationwide clinical trial for children with de novo AML, excluding acute promyelocytic leukemia and myeloid leukemia with Down syndrome, which was conducted between November 2006 and December 2010 in Japan. A central review of diagnosis based on the WHO classification was prospectively performed on each case soon after morphological, cytogenetical and immunological data were submitted to data center. Regarding the cases with discrepant results among these parameters, further diagnostic tests including FISH and chimera gene analyses were underwent to confirm the diagnoses. [Results] Four hundred and eighty four patients were enrolled in the study. Thirty patients did not meet the criteria of AML. We could not collected suitable data for diagnosis in 6 patients. Regarding the rest 448 patients, diagnoses based on the WHO classification 2001 and 2008 were determined. According to the 2001 version, 227 (50.6%) had AML with recurrent genetic abnormalities:124 (27.7%) of AML with t(8;21)(q22;q22);(AML1/ETO ), 32 (7.1%) of AML with inv(16)(p13q22); (CBFβ/MYH11), 38 (8.5%) of AML with t(9;11)(p22;q23), and 33 (7.4%) of AML with the other11q23 (MLL) abnormalities, 36 (8.0%) had AML with multilineage dysplasia, and 185 (41.3%) had AML, not otherwise categorized. According to 2008 version, 235 (52.5%) had AML with recurrent genetic abnormalities: 124 (27.7%) of t(8;21)(q22;q22);(AML1/ETO ), 32 (7.1%) of AML with inv(16)(p13q22); (CBFβ/MYH11), 38 (8.5%) of AML with t(9;11)(p22;q23), 33 (7.4%) of AML with the other11q23 (MLL) abnormalities,4 of AML with t(6;9)(p23;q34);DEK-NUP214,2 of AML with inv(3)(q21q26.2) or t(3;3)(q21;q26.2);RPN1-EVI13, and 2 of AML with t(1;22)(p13;q13);RBM15-MKL, 88 (19.6.7%) had AML with myelodysplasia-related changes (29 from morphological features of myelodysplasia and 59 from myelodysplasia-related cytogenetic abnormalities), 119 (26.6%) had AML, not otherwise categorized and 7(1.6%) had mixed phenotype acute leukemia (6 of T/myeloid and 1 of B/myeloid). [Discussion] Our comprehensive approach for diagnosis was a useful modality for precise diagnosis of uncertain cases, which might have been assigned to the category of AML, with not otherwise categorized, previously. As a result, the present study shows an increased prevalence of AML with recurrent genetic abnormalities or AML with myeloid dysplasia-related changes among pediatric patients with previously untreated AML. Analysis of the AML-05 trial will elucidate the prognostic impact of these categories. Disclosures: No relevant conflicts of interest to declare.

Array CGH Identifies Copy Number Changes In 10% Of 520 MDS Patients With Normal Karyotype: Deletions Encompass The Genes TET2, DNMT3A, ETV6, NF1, RUNX1, and STAG2 and Are Associated With Shorter Survival

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1516-1516
Author(s):  
Claudia Haferlach ◽  
Melanie Zenger ◽  
Marita Staller ◽  
Andreas Roller ◽  
Kathrin Raitner ◽  
...  
Keyword(s):  
Copy Number ◽  
Chromosome Banding ◽  
Array Cgh ◽  
Normal Karyotype ◽  
Employment Equity ◽  
Interphase Fish ◽  
Banding Analysis ◽  
Chromosome Banding Analysis ◽  
Equity Ownership ◽  
Copy Number Changes

Abstract Background In MDS, cytogenetic aberrations play an important role for classification and prognostication. The original IPSS and the revised IPSS classifiers have clearly demonstrated the prognostic impact of distinct cytogenetic abnormalities. The vast majority of chromosome aberrations in MDS are gains or losses of chromosomal material while balanced rearrangements are rare. However, more than 50% of MDS and even more in low risk MDS harbor a normal karyotype. Chromosome banding analysis can only detect gains and losses of more than 10 Mb size due to its limited resolution and is dependent on proliferation of the MDS clone in vitro to obtain metaphases. Array CGH has a considerably higher resolution and does not rely on proliferating cells. Aims In this study we addressed the question whether MDS with normal karyotype harbor cytogenetically cryptic gains and losses. Patients and Methods 520 MDS patients with normal karyotype were analyzed by array CGH (Human CGH 12x270K Whole-Genome Tiling Array, Roche NimbleGen, Madison, WI). For all patients cytomorphology and chromosome banding analysis had been performed in our laboratory. The cohort comprised the following MDS subtypes: RA (n=22), RARS (n=43), RARS-T (n=27), RCMD (n=124), RCMD-RS (n=111), RAEB-1 (n=104), and RAEB-2 (n=89). Median age was 72.2 years (range: 8.9-90.1 years). Subsequently, recurrently deleted regions detected by array CGH were validated using interphase-FISH. Results In 52/520 (10.0%) patients copy number changes were identified by array CGH. Only eight cases (1.5%) harbored large copy number alterations >10 Mb in size, as such generally detectable by chromosome banding analysis. These copy number alterations were confirmed by interphase-FISH. They were missed by chromosome banding analysis due to small clone size (n=2), insufficient in vitro proliferation (n=3) or poor chromosome morphology (n=3). In the other 44 patients with submicroscopic copy number alterations 18 gains and 32 losses were detected. The sizes ranged from 193,879 bp to 1,690,880 bp (median: 960,176 bp) in gained regions and 135,309 bp to 3,468,165 bp (median: 850,803 bp) in lost regions. Recurrently deleted regions as confirmed by interphase-FISH encompassed the genes TET2 (4q24; n=9), DNMT3A (2p23; n=3), ETV6 (12p13; n=2), NF1 (17q11; n=2), RUNX1 (21q22; n=2), and STAG2 (Xq25, deleted in 2 female patients). No recurrent submicroscopic gain was detected. In addition, we performed survival analysis and compared the outcome of patients with normal karyotype also proven by array CGH (n=462) to patients with aberrant karyotype as demonstrated by array CGH (n=52). No differences in overall survival were observed. However, overall survival in 35 patients harboring deletions detected solely by array CGH was significantly shorter compared to all others (median OS: 62.1 vs 42.4 months, p=0.023). Conclusions 1. Array CGH detected copy number changes in 10.0% of MDS patients with cytogenetically normal karyotype as investigated by the gold standard method, i.e. chromosome banding analysis. 2. Most of these alterations were submicroscopic deletions encompassing the genes TET2, ETV6, DNMT3A, NF1, RUNX1, and STAG2. 3. Interphase-FISH for these loci can reliably pick up these alterations and is an option to be easily performed in routine diagnostics in MDS with normal karyotype. 4. Patients harboring deletions detected solely by array-CGH showed worse prognosis. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Zenger:MLL Munich Leukemia Laboratory: Employment. Staller:MLL Munich Leukemia Laboratory: Employment. Roller:MLL Munich Leukemia Laboratory: Employment. Raitner:MLL Munich Leukemia Laboratory: Employment. Holzwarth:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kohlmann:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

scholarly journals Multiplex ligation-dependent probe amplification identifies copy number changes in normal and undetectable karyotype MDS patients

2020 ◽  
Author(s):  
Jing Ma ◽  
xiaofei Ai ◽  
Jinhuan Wang ◽  
Limin Xing ◽  
Chen Tian ◽  
...  
Keyword(s):  
Copy Number ◽  
Chromosome Banding ◽  
Chromosomal Abnormalities ◽  
Normal Karyotype ◽  
Prognostic Information ◽  
Significant Difference ◽  
Banding Analysis ◽  
Chromosome Banding Analysis ◽  
Copy Number Changes ◽  
Dependent Probe

Abstract Background Chromosomal abnormalities play an important role in classification and prognostication of myelodysplastic syndromes (MDS) patients. However, more than 50% low risk MDS patients harbor a normal karyotype. Recently, multiplex ligation-dependent probe amplification (MLPA) has emerged as an effective and robust method for the detection of cytogenetic aberrations in MDS patients. Methods To characterize the subset of MDS with normal karyotype or failed chromosome banding analysis, we analyzed 144 patient samples with normal karyotype or undetectable through regular chromosome banding, which were subjected to parallel comparison via fluorescence in situ hybridization (FISH) and MLPA. Results MLPA identifies copy number changes in 16.7% of 144 MDS patients and we observed a significant difference in overall survival (OS) (median OS: undefined vs 27 months, p=0.0071) in patients with normal karyotype proved by MLPA, versus aberrant karyotype cohort as determined by MLPA. Interestingly, patients with undetectable karyotype via regular chromosome banding indicated inferior outcome. Conclusion Collectively, MDS patients with normal or undetectable karyotype via chromosome banding analysis can be further clarified by MLPA, providing more prognostic information that benefit for individualized therapy.

scholarly journals Molecular Cytogenetic Delineation of Deletions and Translocations Involving Chromosome Band 7q22 in Myeloid Leukemias

Blood ◽  
1997 ◽  
Vol 89 (6) ◽  
pp. 2036-2041 ◽  
Author(s):  
Konstanze Fischer ◽  
Stefan Fröhling ◽  
Stephen W. Scherer ◽  
Jill McAllister Brown ◽  
Claudia Scholl ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Myeloid Leukemia ◽  
Chromosome Banding ◽  
De Novo ◽  
Chromosome Band ◽  
Reciprocal Translocations ◽  
Myeloid Leukemias ◽  
Banding Analysis ◽  
Chromosome Banding Analysis ◽  
Translocation Breakpoints

Abstract Loss of chromosome 7 (−7) or deletion of its long arm (7q−) are recurring chromosome abnormalities in myeloid disorders, especially in therapy-related myelodysplastic syndrome (t-MDS) and acute myeloid leukemia (t-AML). The association of −7/7q− with myeloid leukemia suggests that these regions contain a novel tumor suppressor gene(s) whose loss of function contributes to leukemic transformation or tumor progression. Based on chromosome banding analysis, two critical regions have been identified: one in band 7q22 and a second in bands 7q32-q35. We analyzed bone marrow and blood samples from 21 patients with myeloid leukemia (chronic myeloid leukemia, n = 2; de novo MDS, n = 4; de novo AML, n = 13; t-AML, n = 2) that on chromosome banding analysis exhibited deletions (n = 19) or reciprocal translocations (n = 2) of band 7q22 using fluorescence in situ hybridization. As probes, we used Alu-polymerase chain reaction products from 22 yeast artificial chromosome (YAC) clones that span chromosome bands 7q21.1-q32, including representative clones from a panel of YACs recognizing a contiguous genomic DNA fragment of 5 to 6 Mb in band 7q22. In the 19 cases with deletions, we identified two distinct commonly deleted regions: one region within band 7q22 was defined by the two CML cases; the second region encompassed a distal part of band 7q22 and the entire band 7q31 and was defined by the MDS/AML cases. The breakpoint of one of the reciprocal translocations was mapped to 7q21.3, which is centromeric to both of the commonly deleted regions. The breakpoint of the second translocation, which was present in unstimulated bone marrow and phytohemagglutinin-stimulated blood of an MDS patient, was localized to a 400-kb genomic segment in 7q22 within the deletion cluster of the MDS/AML cases. In conclusion, our data show marked heterogeneity of 7q22 deletion and translocation breakpoints in myeloid leukemias, suggesting the existence of more than one pathogenetically relevant gene.

AML with Inv(3)(q21q26) or t(3;3)(q21;q26) Are Frequently Accompanied by Mutations in RUNX1 and NRAS and Show a High Incidence of NF1 Deletions: a Study On 40 Cases.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 273-273
Author(s):  
Claudia Haferlach, ◽  
Torsten Haferlach ◽  
Frank Dicker ◽  
Tamara Weiss ◽  
Wolfgang Kern ◽  
...  
Keyword(s):  
Cell Count ◽  
Chromosome Aberrations ◽  
De Novo ◽  
Blast Cell ◽  
Splice Mutation ◽  
Acute Leukemias ◽  
Genetic Abnormalities ◽  
Additional Chromosome ◽  
Equity Ownership ◽  
De Novo Aml

Abstract Abstract 273 AML with inv(3)(q21q26) or t(3;3)(q21;q26) have been defined as a distinct entity in the WHO classification in the category of “acute myeloid leukemia with recurrent genetic abnormalities”. Whereas cases with t(8;21)(q22;q22), inv(16)(p13q22/t(16;16)(p13;q22) or t(15;17)(q22;q12) are considered as acute leukemias regardless of the percentage of blasts in the bone marrow, it is not clear whether cases with inv(3)(q21q26)/t(3;3)(q21;q26) should be categorized as such if blast cell count is <20%. To analyze the spectrum of myeloid neoplasms in which inv(3)/t(3;3) occurs, all cases with these abnormalities as diagnosed since 2005 in our laboratory were identified. CML cases showing a t(9;22) and inv(3)/t(3;3) were not included as here CML blast crisis was evident. To further decipher accompanying genetic lesions in AML with inv(3)(q21q26)/t(3;3)(q21;q26) we performed in addition to chromosome banding analyses, FISH for the detection of NF1 deletions and mutation screening of NPM1, MLL (PTD), FLT3 (ITD, TKD), RUNX1, KIT (D816), NRAS (codon12/13/61), CBL (exon8/9 splice mutation), MPL, and JAK2 (V617F, exon12). The study cohort included 20 males and 20 females. Median age was 64.8 years (range: 36.3–91.3), median WBC was 3.9G/l (range: 1.1–75.0G/l) and median platelet count 133G/l (range: 5–799G/l). Based on cytomorphology 23 cases were classified as AML (de novo: 18, t-AML: 2, s-AML: 3 (2 after MPN, 1 after MDS), 15 as MDS and 2 as MPN. 27 showed an inv(3)(q21q26) and 13 a t(3;3)(q21;q26). Additional chromosome aberrations were observed in 23/40 (57.5%) cases (AML: 16/23, 69.6%; MDS 7/17, 41.2%): one, two or more than two additional aberrations in 13, 6 and 4 cases, respectively. Recurrent abnormalities were −7 (n=17), del(5q) (n=4). The following molecular mutations were detected: FLT3-TKD (1/32, 3.1%), NRAS (7/29, 24%), RUNX1 (6/29, 20.7%), CBL exon8/9 splice mutation (n=2/24, 8.3%) and JAK2V617F (2/27, 7.4%). One case each with JAK2V617F was diagnosed with MPN and s-AML after MPN, respectively. No mutations were detected for: NPM1 (0/20), MLL-PTD (0/22), FLT3-ITD (0/35), KIT (0/25), JAK2exon12 (0/13) and MPL (0/13). One copy of the NF1 (neurofibromin 1) gene, which negatively regulates the RAS pathway, was found to be deleted in 4/26 (15.4%) cases using FISH (NF1/MPO probe from Kreatech, Amsterdam, The Netherlands). Overall, 22 molecular alterations were observed in the analyzed genes (15 in AML cases, 7 in MDS/MPN cases). Taking also additional cytogenetic aberrations into account 31/40 patients (20/23 AML, 11/17 MDS/MPN) showed further genetic abnormalities in addition to inv(3)/t(3;3). Survival data were available in 28 cases. No significant differences were observed with respect to overall survival (OS) and event-free survival (EFS) between cases diagnosed as MDS or MPN vs AML. Whereas no impact of additional chromosome aberrations or presence of molecular mutations on OS was observed, a trend for a shorter OS in cases with RUNX1 mutation was found (2 months vs 21.8 months, p=0.07). In addition 17 cases with inv(3)(q21q26)/t(3;3)(q21;q26) and de novo AML were compared to 814 de novo AML without inv(3)(q21q26)/t(3;3)(q21;q26). The median OS for the total cohort was 48.2 months (mo), the median EFS 16.3 mo. Based on cytogenetics cases were assigned into 8 subgroups: 1. t(15;17)(q22;q21), 2. t(8;21)(q22;q22), 3. inv(16)(p13q22)/t(16;16)(p13;q22), 4. 11q23/MLL abnormalities, 5. inv(3)(q21q26)/t(3;3)(q21;q26), 6. normal karyotype, 7. complex karyotype, 8. other abnormalities. Median OS was not reached for groups 1, 2, 3, 4, and 6 and was 23.4 mo, 11.8 mo and 32.2 mo for groups 5, 7, and 8, respectively. OS at 2 yrs was 95.6%, 96.3%, 76.6%, 64.9%, 47.5%, 63.5%, 23.9% and 58.5% for groups 1–8, respectively. The respective data for median EFS were: not reached for groups 1 and 2 and 15.9 mo, 13.5 mo, 6.3 mo, 16.9 mo, 7.5 mo and 12.5 mo for groups 3–8, respectively. In summary, inv(3)/t(3;3) is observed in MDS, MPN, de novo AML, s-AML and t-AML and frequently accompanied by additional cytogenetic or molecular genetic abnormalities. Especially frequent were mutations in RUNX1, NRAS and deletions of NF1. Prognosis of patients with inv(3)/t(3;3) is unfavourable irrespective of the cytomorphological diagnosis. These data suggest to consider cases with inv(3)/t(3;3) as one entity regardless of blast cell count. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Equity Ownership. Dicker:MLL Munich Leukemia Laboratory: Employment. Weiss:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Equity Ownership.

IDH1/2, TET2 and DNMT3A Mutations Are Not Mutually Exclusive in Secondary Acute Myeloid Leukemias,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3556-3556
Author(s):  
Olivier Kosmider ◽  
Olivier LaRochelle ◽  
Marie-Magdelaine Coude ◽  
Veronique Mansat-De Mas ◽  
Eric Delabesse ◽  
...  
Keyword(s):  
Bone Marrow ◽  
De Novo ◽  
Conflicts Of Interest ◽  
Remission Rate ◽  
Normal Karyotype ◽  
Prognostic Impact ◽  
Mutational Status ◽  
Myeloid Leukemias ◽  
Melting Analysis ◽  
De Novo Aml

Abstract Abstract 3556 IDH1/2, TET2 and DNMT3A mutations have been reported in myeloid malignancies including de novo AML. In this study, we have analyzed the frequency and prognostic impact of these mutations in a large retrospective cohort of patients (pts) with secondary AML (SA) which encompass myelodysplasia-related changes (MRC) AML and therapy-related (TR) AML according to the WHO classification. Bone marrow samples were collected from 247 pts at diagnosis with SA and the mutational status of IDH1/2, TET2 and DNMT3A genes together with other genes frequently mutated in AML (NPM1, FLT-3, N and K-RAS, WT1) was determined by Sanger sequencing or high resolution melting analysis. The cohort of 247 pts consisted in 201 MRC AML and in 46 TR AML, 39.5% of which with a normal karyotype (NK). The frequency of IDH1/2, TET2 and DNMT3A mutations was 12.6, 19.8 and 4.5%, respectively. Two pts had both TET2 and IDH1/2 mutations, 2 pts had TET2 and DNMT3A mutations and 5 pts had both IDH1/2 and DNMT3A mutations showing that these mutations were not mutually exclusive in SA. IDH1/2 and TET2 mutations were significantly more frequent in MRC AML (14.1 and 22.3%) than in TR AML (6.4 and 8.7%) (P =0.04 and P =0.03) while the frequency of DNMT3A mutations was identical in the two subgroups. The SA pts harbouring at least one IDH1/2 or TET2 or DNMT3A mutation were significantly older (P <0.0001) and presented higher leukocyte count and lower MCV (P <0.05) than unmutated pts. Percentage of blasts in the bone marrow was similar in the two groups. Karyotype was normal in 48% of the IDH1/2 or TET2 or DNMT3A mutated pts and 18% of the unmutated patients, indicating that these mutations were strongly associated with NK (P < 0.001). A statistically significant link was found between TET2 or IDH1/2 or DNMT3A mutations and NPM1 mutations, but not with FLT-3, N/K-RAS or WT1 mutations. Complete remission rate and overall survival were evaluated in a group of 158 pts which had received intensive chemotherapy at diagnosis, and were identical in the IDH1/2 or TET2 or DNMT3A mutated and unmutated groups. These mutations did significantly influence survival neither in the subgroup of pts with normal karyotype, nor in the subgroup of MRC-AML, or TR-AML which were of very poor prognosis. These data show that IDH1/2, TET2 or DNMT3A mutations could modify the clinical presentation without impact on prognosis. Disclosures: No relevant conflicts of interest to declare.

AML with 11q23/MLL abnormalities as defined by the WHO classification: incidence, partner chromosomes, FAB subtype, age distribution, and prognostic impact in an unselected series of 1897 cytogenetically analyzed AML cases

Blood ◽  
2003 ◽  
Vol 102 (7) ◽  
pp. 2395-2402 ◽  
Author(s):  
Claudia Schoch ◽  
Susanne Schnittger ◽  
Mirjam Klaus ◽  
Wolfgang Kern ◽  
Wolfgang Hiddemann ◽  
...  
Keyword(s):  
Who Classification ◽  
De Novo ◽  
Age Distribution ◽  
Prognostic Impact ◽  
Monocytic Differentiation ◽  
Dismal Prognosis ◽  
Genetic Abnormalities ◽  
De Novo Aml ◽  
11Q23 Abnormalities ◽  
French American British

Abstract Acute myeloid leukemia (AML) cases with 11q23 abnormalities involving the MLL gene comprise one category of recurring genetic abnormalities in the WHO classification. In an unselected series of 1897 AML cases, 54 patients with an 11q23/MLL rearrangement were identified, resulting in an incidence of 2.8%. The incidence of AML with MLL rearrangement was significantly higher in therapy-related AML (t-AML) than in de novo AML (9.4% vs 2.6%, P &lt; .0001). The frequency of MLL rearrangements was significantly higher in patients younger than 60 years (5.3% vs 0.8%, P &lt; .0001). While the incidence of MLL rearrangements in AML M4, M5a, and M5b was 4.7%, 33.3%, and 15.9%, respectively, it was found in only 0.9% of all other French-American-British (FAB) subtypes (P &lt; .0001). Compared with AML with intermediate karyotype, AML with 11q23/MLL rearrangement had a worse outcome, which was rather comparable with AML with unfavorable karyotype. Compared with t-AML, the median overall survival (OS) of de novo AML with MLL rearrangement was significantly better (2.5 vs 10 months, P = .0143). No significant differences in median OS were observed between cases with t(9;11) compared with all other MLL rearrangements (10.0 vs 8.9 months, P = .36). In conclusion, the category AML with 11q23/MLL abnormalities accounts for 2.8% of unselected AML, is closely associated with monocytic differentiation, and has a dismal prognosis. (Blood. 2003;102:2395-2402)

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