Mutations In SETBP1 Occur In 3.1% Of De Novo AML and Show a Distinct Genetic Pattern That Highly Resembles Atypical CML

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2560-2560
Author(s):  
Manja Meggendorfer ◽  
Tamara Alpermann ◽  
Elisabeth Sirch ◽  
Claudia Haferlach ◽  
Wolfgang Kern ◽  
...  
Keyword(s):  
Sanger Sequencing ◽  
De Novo ◽  
Complex Karyotype ◽  
Employment Equity ◽  
Cytogenetic Abnormalities ◽  
Myeloid Malignancies ◽  
Genetic Pattern ◽  
Equity Ownership ◽  
De Novo Aml ◽  
Atypical Cml

Abstract Introduction Recently, mutations in SETBP1 (SETBP1mut) have been identified in different myeloid malignancies. We previously determined mutation frequencies in the range of 5-10% in MPN and MDS/MPN overlap, while we found SETBP1 more frequently mutated in atypical CML (32%). SETBP1mut has been shown to associate with CBL and ASXL1 mutations, as well as the cytogenetic abnormalities -7 and i(17)(q10). While SETBP1 mutations have been detected in 3% of s-AML cases, so far no mutations of SETBP1 in de novo AML have been described. Aim To analyze the mutation frequency of SETBP1 mutations in de novo AML with corresponding cytogenetic abnormalities and their respective correlation to clinical data and other gene mutations. Patients and Methods We investigated 422 adult de novo AML patients, diagnosed by cytomorphology, immunophenotyping and genetic studies following WHO classification. SETBP1 was analyzed by Sanger sequencing of the coding region for amino acids 800 to 935. The cohort comprised 229 males and 193 females, the median age was 65.8 years (range: 19.3 – 89.0). Cytogenetics was available in all 422 cases. Based on the previously described association of SETBP1mut with -7 and i(17)(q10) in other myeloid malignancies there was a selection bias to these karyotypes. Cases were grouped according to cytogenetic abnormalities: normal karyotype (n=88) and aberrant karyotype (n=334), i.e. i(17)(q10) (n=15), +14 (n=20), -7 (n=100), other abnormalities (n=129), and complex karyotype (n=114; 44 contained i(17)(q10), +14 or -7). Within the SETBP1mut cases the following molecular markers were analyzed: ASXL1, CBL, CEBPA, FLT3-ITD, FLT3-TKD, IDH1/2, KRAS, NRAS, NPM1, MLL-PTD, RUNX1, SRSF2, TP53 and WT1 by Sanger sequencing, next generation sequencing, gene scan or melting curve analyses. These data were also available in sub-cohorts of SETBP1 negative cases. Results In the total cohort mutations in SETBP1 were detected in 3.1% (13/422) of all cases. SETBP1mut patients were older (median age: 73.5 vs. 65.7 years; p=0.05) and showed a slightly higher white blood cell count (14.5 vs. 13.8x109/L; p<0.001). There was no correlation to gender, hemoglobin level and platelet count. However, analyzing the cytogenetic groups SETBP1mut showed, like in other myeloid malignancies, a strong co-occurrence with -7 and i(17)(q10), since 4/13 SETBP1 positive cases carried a monosomy 7, and 7/13 an i(17)(q10). The two remaining cases showed a trisomy 14 or a complex karyotype that also contained a i(17)(q10). No SETBP1mut was found in any other cytogenetic subgroup. Therefore, SETBP1mut correlated significantly with i(17)(q10) (8/15 i(17)(q10) were SETBP1mut vs. 5/407 in all other karyotypes; p<0.001). Further, we analyzed the association of SETBP1 mutations with other molecular markers. SETBP1mut correlated with ASXL1mut, 9/33 (27%) ASXL1mut patients showed a mutation in SETBP1, while only 2 (1%) showed a SETBP1 mutation in 229 ASXL1 wild type (wt) patients (p<0.001). This was also true for CBLmut, where 4/8 (50%) CBLmut cases were SETBP1mut, while only 8/158 (5%) were SETBP1mut in the group of CBLwt (p=0.001). This was even more prominent in SRSF2mut patients, where all 9 SRSF2mut were also SETBP1mut, while only 4/8 (50%) patients carried a SETBP1 mutation within the SRSF2wt group (p=0.029). In contrast, SETBP1mut were mutually exclusive of mutations in TP53 (0/67 in TP53mut vs. 12/194 in TP53wt; p=0.04), possibly reflecting the exclusiveness of TP53mut in i(17)(q10) patients. There was no correlation to any other analyzed gene mutation. Remarkably, while there was a high coincidence of SETBP1mut, SRSF2mut (9/13) and ASXL1mut (9/11), none of these patients showed mutations in the typical AML markers NPM1, FLT3-ITD, CEBPA, MLL-PTD, or WT1. Comparing the mutational loads of SETBP1, ASXL1 and SRSF2 resulted in SRSF2 having in most cases the highest mutational loads (range: 30-70%) while ASXL1 and SETBP1 showed equal or lower mutational loads (15-50% and 10-50%, respectively), possibly indicating that SRSF2 mutation is a former event followed by ASXL1 and SETBP1 mutation. Conclusions 1) For the first time we describe, that SETBP1 mutations occur in de novo AML. 2) SETBP1 mutations are correlated with a distinct genetic pattern with high association to i(17)(q10), ASXL1 and SRSF2 mutations and are mutually exclusive of TP53mut. 3) Thus, the genetic pattern of SETBP1 mutated AML highly resembles that of atypical CML. Disclosures: Meggendorfer: MLL Munich Leukemia Laboratory: Employment. Alpermann:MLL Munich Leukemia Laboratory: Employment. Sirch: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.

Myeloid Malignancies with Isolated 7q Deletion Can be Further Characterized By Their Accompanying Molecular Mutations

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3811-3811
Author(s):  
Claudia Haferlach ◽  
Annette Fasan ◽  
Manja Meggendorfer ◽  
Melanie Zenger ◽  
Susanne Schnittger ◽  
...  
Keyword(s):  
Array Cgh ◽  
De Novo ◽  
Jak2 V617f ◽  
Terminal Deletion ◽  
Employment Equity ◽  
Myeloid Malignancies ◽  
Additional Chromosome ◽  
Equity Ownership ◽  
De Novo Aml ◽  
7Q Deletion

Abstract Background: 7q deletions (del(7q)) are recurrent cytogenetic abnormalities. They occur either as the sole abnormality or accompanied by additional chromosome aberrations in AML, MDS, MDS/MPN and MPN. Cases with del(7q) as the sole abnormality are rare and poorly characterized. Aim: In patients with myeloid malignancies and del(7q) as the sole abnormality we determined 1. Type and size of the del(7q) 2. Spectrum of accompanying molecular mutations and their impact on the phenotype. Patients and Methods: 81 cases with myeloid malignancies and del(7q) as the sole abnormality were included in this study. Of these 38 had AML (27 de novo, 7 secondary, 4 therapy-related), 17 MDS (14 de novo, 3 therapy-related), 10 MDS/MPN (9 CMML, 1 MDS/MPN unclassifiable) and 16 MPN. The median age was 72 years (range: 29-89 years). All cases were investigated by array CGH (Agilent, Waldbronn, Germany) and for mutations in ASXL1, CALR, CBL, DNMT3A, ETV6, EZH2, JAK2, KRAS, MPL, NPM1, NRAS, RUNX1, SF3B1, SRSF2, TET2, and TP53. Results: Array CGH revealed an interstitial del(7q) in 67 cases, while 14 cases showed terminal del(7q). Further characterization of these deletions using 24 color FISH revealed unbalanced translocations in 10 of the 14 cases with terminal deletion. Partner chromosomes were X, 8, 9, 12, 13, 17 (n=2), 19 (n=2), and 22. The breakpoints on chromosome 7 were diverse ranging from 7q11 to 7q32. In two cases the breakpoint was within the CDK6 gene. In two cases with terminal del(7q) the complete loss of 7q was due to an idic(7)(q11.21). In the remaining two cases the terminal deletion could not be further resolved. In the 67 cases with interstitial del(7q) the size of the del(7q) varied between 1.8 and 158.9 Mb (median: 52.6 Mb). No commonly deleted region could be identified for all cases. However, in 57 cases the deleted region encompassed genomic position 101,912.442 (7q22.1) to 119,608.824 (7q31.31) including 111 genes. The size of the 7q deletion was smaller in cases with interstitial deletion as compared to terminal deletion (57.7 MB vs 70.9 MB, p=0.04) and in MPN as compared to all other entities (48.7 MB vs 62.8 MB, p<0.001). The mutation analyses revealed mutations in TET2 37% (25/67), ASXL1 35% (27/78), RUNX1 26% (18/69), DNMT3A 21% (14/68), SRSF2 18% (13/73), JAK2 V617F 14% (11/79), CBL 9% (7/75), NRAS 9% (7/77), MLL -PTD 5% (4/80), KRAS 5% (3/66), EZH2 4% (3/72), TP53 4% (3/74), SF3B1 4% (3/75), ETV6 3% (2/73), NPM1 3% (2/77), CALR 1% (1/77), MPL 1% (1/76). ASXL1 and TET2 were frequently co-mutated as 56% of ASXL1 mutated cases also harbored a TET2 mutation (p=0.02). 39 cases were analysed for all 16 molecular mutations. The majority of patients (n=27, 69%) had more than one mutation (range: 2-4), 9 patients (23%) had one mutation and in 3 patients (8%) no mutation was detected. The number of mutations per patient was lower in patients <70 years as compared to patients ≥70 years (0, 1,2,3,4 mutations detected in: 23%, 15%, 15%, 46%, and 0% vs 0%, 27%, 27%, 31%, and 15%, p=0.05). CBL mutations were most frequent in CMML (44%) but rare in all other subtypes (5%, p=0.003), while RUNX1 mutations were most frequent in AML (43% vs 9%; p=0.002) and JAK2 V617F mutations most frequent in MPN (50% vs 5%, p<0.001). DNMT3A mutations and MLL -PTD were significantly more frequent in de novo AML than in all other entities (43% vs 11%, p=0.007; 15% vs 0%, p=0.009), while no significant differences in frequency were observed between the different entities for any of the other mutations or the number of mutations per case. In CMML CBL mutations were associated with del(7q) (44%) as CBL mutations were present in only 17% of non del(7q) CMML (n=101, p=0.07). The frequency of RUNX1 mutations was significantly higher in AML with del(7q) as the sole abnormality (43%) as compared to all other AML (n=2273, 21%; p=0.001). Median overall survival (OS) for the total cohort was 25 months and did not differ significantly between AML, MDS, MDS/MPN and MPN (26, 27, not reached, 15 months, respectively). Conclusions: 1. Sizes and localisations of the del(7q) largely overlapped between AML, MDS, MDS/MPN and MPN. 2. 92% of all patients with 7q deletion harbored at least 1 molecular mutation. 3. TET2 and ASXL1 were the most frequently mutated genes and were present at comparable frequencies in all subtypes. 4. AML with del(7q) is closely associated with RUNX1 mutations while CMML with del(7q) frequently harbored CBL mutations suggesting a cooperative leukemogenic potential in these entities. Disclosures Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Fasan:MLL Munich Leukemia Laboratory: Employment. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Zenger: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.

ETV6 Rearrangements Are Recurrent Genetic Events In Myeloid Malignancies and In AML Are Associated with NPM1- and RUNX1-Mutations and An Intermediate Prognosis

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2758-2758
Author(s):  
Claudia Haferlach ◽  
Susanne Schnittger ◽  
Wolfgang Kern ◽  
Torsten Haferlach
Keyword(s):  
De Novo ◽  
Normal Karyotype ◽  
Employment Equity ◽  
Childhood All ◽  
Myeloid Malignancies ◽  
Equity Ownership ◽  
De Novo Aml ◽  
Diagnostic Work ◽  
Work Up

Abstract Abstract 2758 Introduction: The ETV6 gene (formerly TEL) is located in the chromosomal band 12p13 and is a frequent target of deletions and chromosomal translocations in both myeloid and lymphoid leukemias. In ALL the most frequent partner gene of ETV6 is RUNX1. ALL with ETV6-RUNX1 fusions are observed in 20% of childhood ALL and are associated with favorable outcome. In contrast ETV6 rearrangements are less frequent and not well described in myeloid malignancies. Therefore, the aim of this study was to analyze ETV6 rearrangements in myeloid malignancies with respect to frequency, partner genes and impact on prognosis. Patients/Methods: 55 cases with ETV6 rearrangements were identified in a total cohort of 9,550 cases (0.5%) with myeloid malignancies (de novo AML: n=3,090, s-AML: 486, t-AML: 222, MDS: n=3,375, MDS/MPN overlap: n=210, CMML: n=447, MPN: n=1,720) which had been sent to our laboratory between 08/2005 and 07/2010 for diagnostic work-up. In all cases chromosome banding analysis was performed and in cases with abnormalities involving 12p13 FISH was carried out in addition to verify the ETV6 rearrangement. Results: ETV6 rearrangements were observed in 31 patients with de novo AML (1.0% of investigated cases), 8 with s-AML (1.7%), 5 with t-AML (2.3%), 6 with MDS (0.2%) and 5 with MPN (0.3%). No ETV6 rearrangements were detected in the cohorts of MDS/MPN or CMML. ETV6 rearrangements were significantly more frequent in s-AML and t-AML as compared to de novo AML (p<0.001). Median age in AML was 59.9 years. In 15 cases with de novo AML FAB-subtypes were available: M0: n=8, M1: n=4, M2: n=1, M4: n=1, and M7: n=1. Thus, ETV6 rearrangements are closely related to immature AML subtypes. In 25/55 cases (45.5%) the ETV6 rearrangement was the sole abnormality. Recurrent additional abnormalities were 7q-/-7 in 10 cases and del(5q) in 8 cases. 36 different partners of ETV6 were observed, recurrent partners were located on 3q26 (EVI1, n=11), 5q33 (PDGFRB, n=4), 22q12 (n=3), 2q31 (n=2), 5q31 (ACSL6, n=2), 12p12 (n=2), 17q11 (n=2). Molecular analysis was performed in addition in AML with ETV6 rearrangements for mutations in NPM1 (n=26 investigated), FLT3-ITD (n=33), FLT3-TKD (n=11), MLL-PTD (n=25) and RUNX1 (n=7). NPM1-mutations were observed in 5 cases (19.2%), FLT3-ITD in 3 cases (9.1%), FLT3-TKD in 2 cases (18.2%), MLL-PTD in 1 case (4%) and RUNX1 mutations in 4 cases (57.1%), respectively. Clinical follow-up data was available of 47 cases. No differences in overall survival (OS) and event-free survival (EFS) were observed in cases with ETV6 rearrangement whether or not additional cytogenetic abnormalities or 7q-/-7 or del(5q) were present. Next 30 de novo AML with ETV6 rearrangement were compared to 819 AML without ETV6 rearrangement. Based on cytogenetics cases were assigned into 9 subgroups: 1) t(15;17)(q22;q21), n=48; 2) t(8;21)(q22;q22), n=29; 3) inv(16)(p13q22)/t(16;16)(p13;q22), n=19; 4) 11q23/MLL abnormalities, n=28; 5) inv(3)(q21q26)/t(3;3)(q21;q26), n=6; 6) normal karyotype, n=424; 7) complex karyotype, n=71; 8) other abnormalities, n=194 and 9) ETV6 rearrangements, n=30. Median OS was not reached for groups 1, 2, 3, 4, and 6 and was 10.6 mo, 11.8 mo, 32.2 and 26.3 mo for groups 5, 7, 8, and 9 respectively. OS at 2 yrs was 95.6%, 96.3%, 76.6%, 64.9%, 26.7%, 63.3%, 23.9%, 58.5% and 60.1% for groups 1–9, respectively. The respective data for median EFS were: not reached for groups 1 and 2 and 15.9 mo, 13.5 mo, 5.1 mo, 16.6 mo, 7.5 mo, 12.5 mo and 14.0 mo for groups 3–9, respectively. Conclusions: ETV6 rearrangements are rare in myeloid malignancies. ETV6 is rearranged with a large variety of partner genes. The highest frequency of ETV6 rearrangements was observed in s-AML and t-AML. OS and EFS of AML with ETV6 rearrangements are comparable to AML with normal karyotype. Thus, the detection of ETV6 rearrangements is associated with in intermediate prognosis. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Myeloid Malignancies With Isochromosome 17q Harbor Frequently Mutations In ASXL1, SETBP1, and SRSF2 - This Distinct Genotype Presents With Various Morphological Phenotypes

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1364-1364
Author(s):  
Manja Meggendorfer ◽  
Tamara Alpermann ◽  
Claudia Haferlach ◽  
Elisabeth Sirch ◽  
Wolfgang Kern ◽  
...  
Keyword(s):  
Clonal Evolution ◽  
Normal Karyotype ◽  
Employment Equity ◽  
Cytogenetic Abnormalities ◽  
Myeloid Malignancies ◽  
Cytogenetic Aberration ◽  
Cytogenetic Aberrations ◽  
Molecular Events ◽  
Equity Ownership ◽  
Atypical Cml

Abstract Introduction The identification of mutations (mut) in SETBP1 recently shed light on a molecular marker in atypical chronic myeloid leukemia (aCML), a disease previously defined by exclusion criteria. SETBP1mut have been identified in different myeloid malignancies. We previously reported mutation frequencies in the range of 5-10% in MPN and MDS/MPN overlap, 32% in aCML, while we found SETBP1 less frequently mutated in AML (3%). SETBP1mut has been shown to associate with ASXL1, CBL and SRSF2 mutations, as well as the cytogenetic abnormalities -7 and i(17)(q10). Aim To investigate the mutation frequency of ASXL1, SETBP1, and SRSF2 in different myeloid entities in correlation to the cytogenetic abnormalities -7 and i(17)(q10). Patients and Methods A cohort of 451 patients (pts) with different myeloid entities was analyzed. Diagnoses according to cytomorphology followed the WHO classification from 2008 (n=439, for n=12 cases no cytomorphology was available): AML (n=29), aCML (n=62), MDS/MPN overlap (n=16), CMML (n=283), MDS (n=5), MPN (n=43), CML (n=1). The cohort consisted of 303 males and 148 females; cytogenetics was available in 445 cases. Patients were grouped by normal karyotype (n=291), i(17)(q10) (n=16), -7 (n=22), and other cytogenetic aberrations (n=117); one case carried both a i(17)(q10) and a -7. ASXL1 exon 13, the mutational hotspot regions of SETBP1 and SRSF2 were analyzed by Sanger sequencing in all cases. Results In the total cohort ASXL1 was mutated in 222/451 (49%), SETBP1 in 61/451 (14%), and SRSF2 in 209/451 (46%) cases. 137 pts showed no mutation in any of these three genes. 171 pts carried one mutation, thereof 84 a sole ASXL1mut, 82 a sole SRSF2mut and only 5 cases showed sole SETBP1mut. In 108 pts two, and in 35 pts all three analyzed genes were mutated. The most frequent combination within the group with two mutations was ASXL1 and SRSF2 (n=78), followed by ASXL1 and SETBP1 (n=16), only 5 cases were mutated in SRSF2 and SETBP1. Addressing the association with cytogenetics revealed that in cases with only one mutation SRSF2mut associated as sole mutation with a normal karyotype (68/124 (55%) SRSF2mut in the normal karyotype group vs. 12/42 (28%) SRSF2mut in all other karyotypes; p=0.003). In contrast, ASXL1mut and SETBP1mut as sole mutations showed no correlation to any addressed karyotype. However, addressing the cases with two mutations the combination of SRSF2mut and ASXL1mut correlated with a normal karyotype (67/291 (23%) SRSF2mut/ASXL1mut in the normal karyotype group vs. 19/154 (12%) SRSF2mut/ASXL1mut in all other karyotypes; p=0.008), while SRSF2mut and SETBP1mut occurred more frequently in i(17)(q10) pts (2/16 (13%) SRSF2mut/SETBP1mut in i(17)(q10) vs. 2/429 (1%) SRSF2mut/SETBP1mut in all other karyotypes; p=0.007). Remarkably, cases with mutations in all three analyzed genes (ASXL1mut, SETBP1mut, and SRSF2mut) highly associated with i(17)(q10) and -7. 11 of 16 cases with i(17)(q10) (69%) showed all three mutations (vs. 24/429 (6%) in all other karyotypes; p<0.001). Furthermore, 6 of 22 cases with -7 (27%) showed mutations in all three genes (vs. 29/423 (7%); p=0.005). Therefore, 15 pts carried all three mutated genes as well as i(17)(q10) or -7. Interestingly, there was no case with only i(17)(q10) and no additional mutation, and only one case with i(17)(q10) and only one additional molecular mutation, 4 cases with two additional molecular mutations and 11 cases carrying all three mutations, possibly indicating that i(17)(q10) appear during clonal evolution. Therefore one might assume that this represents a specific genetic phenotype that is driven by the accumulation of molecular events, since addition of SETBP1mut shifts the association from a normal karyotype to i(17)(q10) or -7. Analyzing the distribution of these cases for mutations in all three analyzed genes +/- additional cytogenetic aberration i(17)(q10) or -7 in the different myeloid entities showed that AML, aCML, CMML, MDS as well as MPN showed this genetic phenotype (AML: n=7 (24%); atypical CML: n=12 (19%); CMML: n=8 (3%); MDS: n=1 (20%); MPN: n=6 (14%)). Conclusions Mutations in SETBP1 associate with ASXL1mut and SRSF2mut and are frequently found in patients with i(17)(q10) or -7. This combination of genetic lesions occurs in different myeloid entities and might therefore define a specific genetically defined subtype of myeloid malignancy. Disclosures: Meggendorfer: MLL Munich Leukemia Laboratory: Employment. Alpermann:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Sirch: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.

RUNX1 Mutated AML Are Associated with a Distinct Pattern of Cytogenetic and Additional Molecular Genetic Abnormalities

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 124-124
Author(s):  
Niroshan Nadarajah ◽  
Susanne Schnittger ◽  
Wolfgang Kern ◽  
Torsten Haferlach ◽  
Claudia Haferlach
Keyword(s):  
High Frequency ◽  
De Novo ◽  
Male Gender ◽  
The Other ◽  
Typical Pattern ◽  
Employment Equity ◽  
Cytogenetic Abnormalities ◽  
Genetic Abnormalities ◽  
Equity Ownership ◽  
De Novo Aml

Abstract Background: Mutations in RUNX1 have been reported in 5 to 20% of AML. The aim of this study was to analyse a large cohort of RUNX1 mutated AML in detail with respect to accompanying cytogenetic and moleculargenetic abnormalities, mutation type and mutation load. Patients and Methods: We investigated 468 AML with RUNX1 mutations (mut) all identified during diagnostic work-up in our laboratory. Sequencing was performed by either Sanger or next-generation sequencing. Median age was 71.9 yrs (range 18-91 yrs), male : female ratio 297 : 171. 369 patients had de novo AML, 75 s-AML following MDS, 24 t-AML. For all patients (pts) cytogenetics was available and categorized according to MRC (Grimwade et al. Blood 2010). Mutation data was available for NPM1 (n=455), MLL-PTD (n=454), CEBPA (n=449), FLT3-TKD (n=422), WT1 (n=394), FLT3-ITD (n=362), ASXL1 (n=293), TP53 (n=204), DNMT3A (n=143), TET2 (n=143), and SF3B1 (n=99). Data on FAB subtype was available in 342 cases (73.1%). Results: The most frequent FAB subtype in RUNX1mut AML was AML M2 (145/342; 42.4%), followed by M1 (23.1%), M4 (15.8%), M0 (15.5%), other subtypes were rare (<2%). Cytogenetics were favorable in 2 (0.4%), intermediate in 397 (84.8%) and adverse in 69 (14.7%) pts. Most frequent cytogenetic abnormalities were +8 (76; 34.7%), +13 (46; 21.0%), +21 (15; 6.8%), +11 (12; 5.5%), -7 (22; 10%), del(7q) (18; 8.2%), del(5q) (12; 5.5%). Only 12 pts (5.5%) showed a complex karyotype (> 3 aberrations). The frequency of all other abnormalities was <5%. Two pts showed a t(15;17)(q24;q21) and 3 MLL rearrangements (partner genes on 7q32, 17q21, 19p13). None of the other recurrent cytogenetic abnormalities according to WHO classification was present. 249 (53.2%) pts had a normal karyotype. ASXL1mut were the most frequent accompanying mutations (36.5%). Mutation frequencies for the other genes were: SF3B1 (27.3%), TET2 (26.6%), FLT3-ITD (17.7%), DNMT3A (16.1%), MLL-PTD (10.8%), CEBPA (6.9%; single mutated (sm): 5.3%, double mutated (dm): 1.6%), WT1 (5.8%), FLT3-TKD (5.5%), TP53 (4.4%), and NPM1 (1.1%). While mutations in ASXL1 and TET2 frequently occurred concomitantly (37.5% of ASXL1mut cases also harboured a TET2mut, p=0.032), ASXL1mut and SF3B1mut rarely co-occurred (only 2 ASXL1mut were SF3B1mut, p=0.001). There were no differences in mutation frequencies and cytogenetic abnormalities observed between de novo AML, s-AML and t-AML. In 368 cases (78.6%) one RUNX1 mutation was detected, 81 pts (17.3%) showed two, 16 cases (3.4%) three, and 3 cases (0.6%) four. The difference in mutation loads between the 2 mutations with the highest load was ≤10% in 55/100 pts, suggesting that both mutations were present in the same clone. In total 592 RUNX1 mutations were detected, 242 (40.9%) were frameshift, 206 (34.8%) missense, 83 (14.0%) nonsense, 37 (6.3%) in frame insertion/deletions, 23 (3.9%) splice-site and 1 no stop change. No association between the types of 1st and 2nd mutations was observed. The mutations were homozygous in 65 pts (13.9%), these were predominantly missense mutations (38; 58.5%). A significantly higher frequency of homozygous mutations was observed in pts with +13 (28.3%, p=0.006), +21 (46.7%; p=0.002), AML M0 (35.8%; p<0.001) and M1 (22.8%; p=0.014) while they were less frequent in M2 (4.8%; p<0.001) and M4 (3.7%, p=0.017). Survival analyses were restricted to 203 de novo AML pts who were treated with intensive chemotherapy (median overall survival (OS) 20.4 months (mo)). Median OS was significantly longer in female than in male pts (45.6 vs 16.3 mo; p=0.003) and in pts ≤ 60 vs >60 yrs (44.4 vs 16.1 mo; p<0.001). Shorter OS was observed for pts with del(5q) (1.8 vs 21.1 mo; p=0.002) and adverse cytogenetics (12.4 vs 23.3 mo, p=0.016). Including these 4 parameters into a multivariate Cox regression analysis revealed that age, male gender and del(5q) were independently associated with shorter OS (relative risk: 1.8, 1.6, 3.4; p: 0.01, 0.038, 0.028) Conclusions: RUNX1 mutated AML: 1. is associated with a myeloid rather than monocytic differentiation, 2. shows a typical pattern of cytogenetic abnormalities with a high frequency of +8 and +13, 3. has a typical pattern of additional molecular mutations with a high frequency of accompanying ASXL1 und SF3B1 mutations, 4. is nearly mutually exclusive of NPM1 and CEBPAdm mutations and other entity defining genetic abnormalities, 5. Male gender, age >60 yrs and del(5q) were negative prognostic factors. Disclosures Nadarajah: 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. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

FISH Analysis In 843 Cases with Myeloid Malignancies Revealed TET2 deletions In 6% of Patients Which Were Accompanied by TET2 Mutations In 51% of Cases

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1706-1706
Author(s):  
Claudia Haferlach ◽  
Sandra Wille ◽  
Alexander Kohlmann ◽  
Susanne Schnittger ◽  
Wolfgang Kern ◽  
...  
Keyword(s):  
Copy Number ◽  
De Novo ◽  
Snp Array ◽  
Suppressor Gene ◽  
Employment Equity ◽  
Myeloid Malignancies ◽  
Tet2 Mutation ◽  
Snp Array Analysis ◽  
Equity Ownership ◽  
De Novo Aml

Abstract Abstract 1706 TET2 (tet oncogene family member 2) on chromosome 4q24 was identified as a candidate tumor suppressor gene. Recurring submicroscopic deletions and copy-neutral loss of heterozygosity (CN-LOH) involving 4q in MDS patients detected by SNP microarray analyses suggested TET2 as an interesting candidate gene. Subsequent sequencing studies revealed TET2 mutations in 10–25% of patients with AML, MDS, and MPNs, while a mutation frequency of up to 42% was reported in CMML. Only a subset of studies evaluated both TET2 mutation status and TET2 copy number status, although this might be of pathophysiological and even of prognostic relevance if TET2 functions as a classical tumor suppressor gene. In the majority of studies copy number status was determined by SNP array analysis, although being expensive and time consuming. Here, in order to investigate TET2 deletions in a large cohort of patients an easy to perform FISH assay was developed applying BACs covering the TET2 gene (RP11-351K6 and RP11-16G16; BlueGnome, Cambridge, UK). This assay was validated on samples with TET2 deletions proven by SNP array analysis. With these FISH probes we analyzed 843 cases with myeloid malignancies (404 AML (323 de novo AML, 68 s-AML, 13 t-AML), 166 MDS, 201 CMML, and 72 MPN). Overall 50 (5.9%) cases with TET2 deletion were identified. These included 22 AML (5.0%), in detail 14 de novo AML (4.3%), 6 s-AML (8.8%), and 2 t-AML (15.4%) as well as 15 CMML (7.5%), 9 MDS (5.4%) and 4 MPN (5.6%). Patients with TET2 deletions showed the following karyotypes: normal: n=15, cytogenetically balanced rearrangements involving 4q24: n=3, 4q deletion as the sole abnormality: n=2, complex: n=25, other abnormalities: n=5. In 25/50 (50%) cases the TET2 deletion was cytogenetically cryptic. In patients with complex aberrant karyotype loss of 4q material was due to interstitial deletion in 7/25 cases and due to unbalanced rearrangements in 16/25 cases, while in 2/25 cases chromosomes 4 were normal in chromosome banding analysis. Furthermore, in patients with TET2 deletions mutation analyses was performed for mutations in TET2 (n=37 investigated), RUNX1 (n=13), NPM1 (n=18), JAK2V617F (n=18), CBL (n=36), NRAS (n=17), KRAS (n=36), FLT3-ITD (n=26), FLT3-TKD (n=8), IDH1 (n=9) and MLL-PTD (n=24). Mutations in TET2 were detected in 19/37 cases (51%), in RUNX1 in 1/13 (8%), in JAK2V617F in 6/18 (33.3%), in CBL in 2/36 (5.6%), in NRAS in 1/17 (6%), in KRAS in 1/36 (2.8%) and in NPM1 in 1/18 (6%) cases, whereas no mutations within the other genes analyzed were found. In the cohort of cases with TET2 deletion and concomitantly TET2 mutation (n=19) 10 had a normal karyotype (52.6%), 5 a complex karyotype (26.3%) and 4 had other abnormalities (21.1%). Importantly, in the cohort of CMML, in 10 of 14 cases (71.4%) both a TET2 deletion and TET2 mutation was detected. Overall, TET2 mutations were significantly more frequent in patients with cytogenetically cryptic TET2 deletion as compared to cytogenetically visible 4q deletions (68.2% vs. 26.7%, p=0.020). In addition FISH screening identified 2 cases (one CMML, one t-MDS) which showed a translocation involving the TET2 locus not leading to a deletion of the BAC signals but a separation suggesting a fusion with yet unidentified partner genes. In conclusion, FISH analyses identified TET2 deletions in 6% of myeloid malignancies. 50% of these deletions were submicroscopic and therefore not detectable by chromosome banding analysis. TET2 deletions were accompanied by TET2 mutations in 51% of respective cases. FISH is a reliable and efficient method to determine the copy number state of TET2. Still, the prognostic impact of TET2 deletions with and without additional TET2 mutations in the various myeloid malignancies has to be evaluated in future investigations. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Wille:MLL Munich Leukemia Laboratory: Employment. Kohlmann: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.

Cytogenetic and Molecular Genetic Characterization Of MLL-PTD Positive AML In Comparison To MLL-Translocated AML

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2557-2557 ◽  
Author(s):  
Claudia Haferlach ◽  
Alexander Kohlmann ◽  
Wolfgang Kern ◽  
Torsten Haferlach ◽  
Susanne Schnittger
Keyword(s):  
De Novo ◽  
Molecular Genetic ◽  
Genetic Alterations ◽  
The Other ◽  
Employment Equity ◽  
Cytogenetic Abnormalities ◽  
Mll Gene ◽  
Equity Ownership ◽  
De Novo Aml ◽  
Tandem Duplications

Background Partial tandem duplications within the MLL gene (MLL-PTD) are a recurrent molecular alteration in acute myeloid leukemia (AML). MLL-PTD occurs with a frequency of 6-8% in de novo AML. Data on additional cytogenetic and molecular alterations in MLL-PTD+ AML is scarce. Beside partial tandem duplications within the MLL gene, the MLL gene is also a target of balanced translocations leading to the fusion of MLL with a large variety of partner genes. Aims 1. Evaluate the spectrum of additional cytogenetic and molecular genetic alterations. 2. Analyze whether additional aberrations impact prognosis. 3. Compare the spectrum of additional abnormalities between MLL-PTD+ AML and AML with MLL-translocations. Patients and Methods We selected a cohort of 225 de novo AML patients harboring a MLL-PTD. These were compared to a cohort of 130 de novo AML with MLL-translocation (MLL-t). Mutation screening for the following genes was performed in pts with MLL-PTD and MLL-t, respectively: ASXL1 (132; 85), CEBPA (184; 67), FLT3-ITD (225; 125), FLT3-TKD (208; 112), IDH1R132 (145; 88), IDH2R140 (141; 63), IDH2R172 (137; 73), KRAS (59; 82), NRAS (98; 82), RUNX1 (213; 97), NPM1 (221; 123), TP53 (104; 89), and WT1 (159; 86). EVI1 expression was assessed in 55 MLL-PTD+ pts and in 77 pts with MLL-t. Results The frequency of MLL-PTD and MLL-translocations differed significantly with respect to age. While MLL-PTD were more frequent in elderly pts, MLL-t were more frequent in younger pts (<10 yrs: 0%/2.3%; 10-19 yrs: 0%/3.8%, 20-29 yrs: 0.9%/13.1%, 30-39 yrs: 3.1%/9.2%, 40-49 yrs: 8.9%/23.1%; 50-59 yrs: 11.1%/13.1%; 60-69 yrs: 35.1%/13.8%; 70-79 yrs: 29.3%/13.8%; ≥80 yrs: 11.6%/7.7%; p<0.001). FAB subtype distribution differed significantly between of MLL-PTD+ and MLL-t AML. While in MLL-PTD+ AML most frequently subtypes M1 (33.8%) and M2 (43.6%) were observed, AML with MLL-t most frequently showed M4 (30%) and M5a (23.1%). The most frequent cytogenetic abnormalities in MLL-PTD+ cases were gains of 11q (n=37), followed by 8q (n=14), and 13q (n=7) and losses of 5q (n=14), 7q (n=14) and 17p (n=5). In contrast, in MLL-t patients the most frequent gains were trisomies 6 (n=7) and 8 (n=33), as well as gains of 1q (n=10), 19p (n=10), 19q (n=8) and 21q (n=21). There were many significant differences in co-occurring mutations between MLL-PTD+ and MLL-t: DNMT3Amut: 44.7% vs 0% (p<0.001), FLT3-ITD: 33.3% vs 3.2% (p<0.001), IDH1R132 14.5% vs 0% (p<0.001), IDH2R140: 19.9% vs 0% (p<0.001), IDH2R172 9.5% vs 0% (p=0.005), and RUNX1 25.8% vs 2.1% (p<0.001). On the other hand KRASmut (3.4% vs 23.2%, p=0.001) and NRASmut (8.2% vs 25.6%, p=0.002) were less frequent in MLL-PTD+ as compared to MLL-t AML. TP53 mutations were observed in comparable frequencies (3.8% vs 5.6%). NPM1 mutations were not detected in either entity. The mean EVI1 expression was significantly higher in MLL-t pts compared to MLL-PTD+ pts (167.1+/-259.1vs 0.4 +/- 0.47, p<0.001). Overall, chromosome abnormalities in addition to the MLL alteration were more frequent in MLL-t AML as compared to MLL-PTD+ AML (mean number of alterations: 1.2 vs 0.7, p=0.004). This goes along with more additional molecular mutations in MLL-PTD+ AML as compared to MLL-t AML (mean number of molecular mutations: 1.5 vs 0.6, p=0.004). Overall survival at 5 yrs was comparable in both subgroups (MLL-PTD+: 27.9% vs MLL-t: 39.8%). In both subgroups age was significantly associated with OS (<60 yrs vs ≥60 yrs: MLL-PTD+: 56.9 vs 16.3 months, MLL-t: 47.8 vs 9.7 months, for both p<0.001). Neither in MLL-PTD+ AML nor in MLL-t AML the presence of additional chromosome aberration had an impact on outcome. With respect to molecular mutations only IDH2R140 was significantly associated with shorter OS (HR: 2.2, p=0.007) and IDH2R172 with longer OS (HR: 0.2, p=0.04) in MLL-PTD+ AML. Conclusions Although both MLL-PTD+ and MLL-translocations disrupt the same gene AML harboring one or the other MLL abnormality differ significantly with respect to age distribution, the pattern of additional cytogenetic abnormalities and the frequency of accompanying molecular mutations. MLL-PTD is more frequent in older patients, presents most frequently as FAB M1 and M2, and harbors more additional molecular genetic events and less additional cytogenetic events. However, both AML subtypes are associated with adverse outcome, particularly in elderly patients. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. 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.

Assessment Of Human Equilibrative Nucleoside Transporter 1 (hENT1) – Expression By Flow Cytometry In Acute Myeloid Leukemia and Myelodysplastic Syndromes and Correlation To Clinical Outcome In Acute Myeloid Leukemia Patients Treated With Intensive Chemotherapy

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2582-2582 ◽  
Author(s):  
Frauke Bellos ◽  
Bruce H. Davis ◽  
Naomi B. Culp ◽  
Birgitte Booij ◽  
Susanne Schnittger ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Myeloid Leukemia ◽  
De Novo ◽  
Molecular Genetic ◽  
Newly Diagnosed ◽  
Employment Equity ◽  
Equity Ownership ◽  
De Novo Aml ◽  
Very High ◽  
Acute Myeloid

Abstract Background Nucleoside analogs depend on cellular hENT1 expression for entry into cells and cytotoxic activity. Studies suggest low cellular hENT1 levels correlate with poor response to such chemotherapies in solid tumors, data on AML and MDS is scarce. Aim To examine hENT1 expression by multiparameter flow cytometry (MFC) in newly diagnosed AML and MDS and correlate results to morphologic, cytogenetic (CG) and molecular genetic (MG) findings. To examine hENT1 expression with respect to clinical outcome in AML patients (pts) treated with intensive cytarabine-based chemotherapy (CHT). Methods We studied pts with newly diagnosed AML (n=145) and MDS (n=96), 133/108 male/female, median age 67.3 (AML) and 73.3 years (MDS). CG was done in 130 AML and 86 MDS. Pts included 107 de novo AML, 9 t-AML, 29 s-AML; FAB: 9 M0, 27 M1, 50 M2, 9 M3, 21 M4, 8 M4eo, 7 M5, 14 not classified; by CG (MRC): 21 favorable, 75 intermediate, 34 adverse. 91 were de novo MDS, 5 t-MDS; 1 RARS, 17 RCMD-RS, 37 RCMD, 3 5q- syndrome, 3 RAEB-1, 5 RAEB-2, 1 CMML, 24 not classified; 2 IPSS-R very low, 55 IPSS-R low, 8 IPSS-R intermediate, 8 IPSS-R high, 13 IPSS-R very high. hENT1 expression was quantified by a novel four color intracellular staining assay using monoclonal antibodies against hENT1, CD45, CD64 and myeloperoxidase. Median fluorescence intensities (MFI) of hENT1 were determined in myeloid progenitors (MP), granulocytes (G) and monocytic cells (Mo) and correlated to hENT1 MFI in lymphocytes to derive hENT1 index (index). Results No correlation of index to age, gender, hemoglobin level or counts for blasts, WBC or platelets was detected. In AML, we generally saw higher index by trend in the more favorable prognostic subgroups. M3/t(15;17)/PML-RARA+ displayed higher index in MP than non-M3 AML (4.24 vs 2.56, p<0.001). G index was lower in M0 (3.01) vs M1, M2, M4 and M4eo (5.66, 4.34, 5.35, 4.77; p=0.01, 0.028, 0.004, 0.043, respectively) and in M2 compared to M1 and M4 (4.34. vs 5.66 and 5.35, p=0.01 and 0.033, respectively). M2 showed lower MP index than M5 (2.42 vs 2.99, p=0.016). Considering CG, index in MP was higher in favorable vs intermediate and adverse pts (3.05 vs 2.58 and 2.53, p=0.034 and 0.023, respectively), Mo index was higher ín favorable vs adverse pts (3.17 vs 2.71, p=0.044). By MG, higher index in Mo and G was observed in RUNX1-RUNX1T1+ AML (4/83, 4.32 vs 3.04, p=0.01; 8.16 vs 4.60, p=0.002, respectively). Higher index for MP was found in FLT3-ITD mutated (mut) (18/111; 3.19 vs 2.62, p=0.012), CEPBA mut (4/26, 3.15 vs 2.35, p=0.004) and for Mo in NPM1 mut AML (23/104; 3.72 vs 2.84, p=0.02), whereas lower index for MP was found in RUNX1mut pts (13/65; 2.17 vs 2.59, p=0.031). De novo AML displayed higher MP index than s-AML (2.7 vs 2.28, p=0.008). Using lowest quartile of index for MP (2.1185) as cut-off, AML pts in the MRC intermediate group treated with CHT (n=38) had inferior OS if MP index was below vs above this cut-off (OS at 6 months 63% vs. 95%, p=0.017, median follow up 4.6 months). MDS showed lower Mo and MP index than AML (2.68 vs 2.96, p=0.021, 1.84 vs 2.65, p<0.001, respectively). By IPSS-R, significance was reached for higher index in Mo and MP in very low risk compared to low risk pts (3.39 vs 2.54, p=0.013 and 4.07 vs 1.78, p<0.001, respectively), MP in very low compared to intermediate and high risk pts (4.07 vs 1.95, p=0.004; 4.07 vs 1.76, p=0.002), and MP and G in very low vs very high risk pts (4.07 vs 1.71, p=0.005; 5.86 vs 3.85, p=0.001, respectively). IPSS-R intermediate vs poor and very poor showed lower G index (5.47 vs 3.59, p=0.018 and vs 3.85, p=0.034 respectively). Conclusion AML with genetic and molecular genetic good risk profile had higher hENT1 expression in MP, G and Mo, suggesting a causal mechanism for better response to CHT and better outcome. Consequently, AML with poor risk molecular genetics (RUNX1 mut) showed lower levels of hENT1 in MP. The detection of higher levels in FLT3-ITD mut pts is in line with reportedly good response to CHT, overall worse outcome being mostly due to early relapses. Strikingly, we saw differences in outcome in pts treated with CHT according to hENT1 expression with shorter OS in pts with low index for MP. Higher index in de novo AML than s-AML and MDS may be causal for better response to nucleoside-based CHT in de novo AML. Data for MDS may be interpreted accordingly, lower risk cases showing higher index in MP, G and Mo. Further analyses are needed to explore hENT1 expression in AML and MDS more comprehensively. Disclosures: Bellos: MLL Munich Leukemia Laboratory: Employment. Davis:Trillium Diagnostics, LLC: Equity Ownership. Culp:Trillium Diagnostics, LLC: Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

scholarly journals The RUNX1 Gene Is Altered in 26% of AML Patients Either By Translocation, Mutation, Gain or Deletion

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 123-123
Author(s):  
Claudia Haferlach ◽  
Niroshan Nadarajah ◽  
Wolfgang Kern ◽  
Susanne Schnittger ◽  
Torsten Haferlach
Keyword(s):  
De Novo ◽  
Intermediate Risk ◽  
Missense Mutations ◽  
Employment Equity ◽  
Runx1 Gene ◽  
Genetic Abnormalities ◽  
Partner Gene ◽  
Different Types ◽  
Equity Ownership ◽  
De Novo Aml

Abstract Background: In AML four types of acquired alterations of the RUNX1 gene have been described: 1. translocations involving RUNX1 leading to fusion genes such as RUNX1-RUNX1T1, 2. molecular mutations, 3. amplifications of RUNX1, 4. Partial or complete deletions of the RUNX1 gene. Aim: To determine the frequency of different RUNX1 alterations and to characterize the spectrum of accompanying genetic abnormalities. Patients and Methods: We screened 726 de novo AML patients (pts) for RUNX1 deletions (del) and translocations using a dual color break-apart probe covering the 5' and 3' part of RUNX1 (MetaSystems, Altlussheim, Germany) and in addition evaluated RUNX1 mutations (mut) by Sanger or next-generation amplicon deep-sequencing. Median age was 67 yrs (range: 18 to 100 yrs). For all patients cytogenetics was available and categorized according to MRC criteria (Grimwade et al. Blood 2010). Partial deletions of RUNX1 as detected by FISH were confirmed by array CGH (Agilent Technologies, Santa Clara, CA). Results: In 89/726 pts (12.3%) abnormalities of the RUNX1 gene were detected by FISH: 10 pts (1.4%) showed a deletion encompassing the whole RUNX1 gene while additional 9 pts (1.2%) showed a partial loss of one RUNX1 copy. A gain of one RUNX1 copy was present in 45/726 (6.2%) pts. In 3 pts a gain of the 5' part of RUNX1 was accompanied by a loss of the 3' part while in 2 pts one copy of the 3' part was gained accompanied by a loss of the 5' part. A translocation affecting the RUNX1 gene was detected in 31 pts (4.3%). The partner gene was RUNX1T1 in 29 pts and located on 16q13 and 18p11 in one pt each. One pt with a RUNX1 translocation also showed a 5' RUNX1 deletion. In 110/726 pts (15.2%) a RUNX1mut was detected. Of these, 16 pts showed two and 5 pts three mutations in RUNX1. Thus, in total 136 mutations were detected in 110 pts: 58 (42.6%) were frameshift, 42 (30.9%) missense, 21 (15.4%) nonsense, 9 (6.6%) splice-site and 6 (4.4%) in-frame insertions/deletions. The RUNX1mut was homozygous in 15 pts, these were predominantly missense mutations (9/15; 60%). Within the subset of pts with RUNX1mut 2 harbored an additional RUNX1del and 9 pts a gain of a RUNX1 copy, while no RUNX1 translocation was present. In AML FAB type M0 both RUNX1mut and RUNX1del showed the highest frequencies (33.3% and 14.8%). 48.4% and 45.2% of cases with RUNX1 translocations were FAB type M1 and M2. While RUNX1mut were most frequent in the cytogenetic intermediate risk group (19.1%; favorable: 2.2%, adverse: 9.7%), RUNX1del were most frequent in pts with adverse risk cytogenetics (9.7%; favorable: 1.1%, intermediate: 0.8%). A comparable distribution was observed for a gain of RUNX1 copies (adverse: 19.4%, favorable: 4.5%, intermediate: 2.6%). With respect to additional molecular mutations all types of RUNX1 alterations were mutually exclusive of NPM1mut. Further, the frequency of DNMT3Amut and CEBPAmut was significantly lower in pts with RUNX1 alterations as compared to those without (14.3% vs. 34.3%; p<0.0001 and 6.4% vs. 13.4%; p=0.012). However, some striking differences between the different types of RUNX1 alterations were detected: ASXL1mut were significantly more frequent in pts with RUNX1mut (36.7%) but rather infrequent in pts with RUNX1del, gain and translocation (12.5%, 6.1%, and 6.7%). A comparable association was noticed for SF3B1mut which were frequent in RUNX1mut pts (23.8%) and rather infrequent in pts with RUNX1del, gain and translocations (0%, 10.5%, and 0%). In contrast, pts with either RUNX1del or RUNX1 gains showed a significantly higher TP53mut frequency (66.7% and 35.3%) as compared to RUNX1mut or RUNX1 translocated pts (7.1% and 4.8%). In the total cohort median overall survival (OS) was 18.7 months and differed significantly between the different types of RUNX1 alterations: for RUNX1 translocations, mutations, gains and deletions it was 35.5, 14.1, 12.4 and 4.3 months. Conclusions: 1. The RUNX1 gene is altered in 26% of AML. 2. All types of RUNX1 alterations predominantly occur in AML M0, M1 and M2 and are rare in the remainder AML. 3. They are mutually exclusive of NPM1 mutations and show a negative association with DNMT3A mutations. 4. While RUNX1 mutations were most frequent in patients with intermediate risk cytogenetics, RUNX1 deletions and gains were most frequent in patients with adverse cytogenetics. 5. Outcome differs significantly and is best in patients with RUNX1 translocations and worst in cases with RUNX1 deletions. Disclosures Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Nadarajah:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Influence of Karyotype On Overall Survival in Patients with Higher-Risk Myelodysplastic Syndrome Treated with Azacitidine or a Conventional Care Regimen.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1755-1755 ◽  
Author(s):  
Ghulam J Mufti ◽  
Steven D. Gore ◽  
Valeria Santini ◽  
Pierre Fenaux ◽  
Lewis R. Silverman ◽  
...  
Keyword(s):  
Overall Survival ◽  
Board Of Directors ◽  
Research Funding ◽  
Conventional Care ◽  
Complex Karyotype ◽  
Trisomy 8 ◽  
Employment Equity ◽  
Advisory Committees ◽  
Cytogenetic Abnormalities ◽  
Equity Ownership

Abstract Abstract 1755 Poster Board I-781 Background Karyotypic abnormalities are common in myelodysplastic syndromes (MDS), and specific chromosomal abnormalities are associated with poor prognosis. The phase III AZA-001 study (Lancet Oncol, 2009) showed azacitidine (AZA) prolonged overall survival (OS) regardless of IPSS cytogenetic risk category. This analysis assessed the effects of specific cytogenetic abnormalities on OS in patient (pt) subgroups treated with AZA or a conventional care regimen (CCR). Methods Pts with higher-risk MDS (FAB RAEB, RAEB-t, or CMML and IPSS Int-2 or High) were enrolled and randomized to receive AZA or CCR. CCR comprised 3 treatments: best supportive care only, low-dose ara-C, or induction chemotherapy. Erythropoietins were prohibited. OS was determined in subgroups of pts with del 5/5q-, del 7/7q-, or trisomy 8, each as part of a non-complex karyotype (<3 cytogenetic abnormalities) or as part of a complex karyotype (≥3 cytogenetic abnormalities). OS was also analyzed in pts with combinations of del 5/5q- and/or del 7/7q- as part of non-complex or complex karyotypes (Table). Pt karyotype was determined at baseline. OS was assessed using Kaplan-Meier methods. A stratified Cox proportional hazards regression model was used to estimate hazard ratios (HRs) and associated 95% confidence intervals (CI). Results A total of 358 pts were enrolled (AZA 179, CCR 179). Of them, 153 had normal karyotypes (AZA 77, CCR 76). Median OS in pts with normal karyotypes was not reached at 21.1 months with AZA vs 17.2 months (95%CI: 15.2 – 24.1 months) with CCR; HR = 0.63 (95%CI: 0.39 – 1.03). Of remaining pts, 136 had del 5/5q-, del 7/7q-, and/or trisomy 8 as part of a non-complex or complex karyotype. AZA was associated with longer OS vs CCR in all subgroups of pts with non-complex cytogenetics, with HRs ranging from 0.20 (95%CI: 0.06 – 0.65) to 0.51 (95%CI: 0.05 – 4.74) (Table). In both the AZA and CCR treatment groups, pts in all subgroups with non-complex karyotypes had substantially longer OS than pts with complex karyotypes. Pts with complex karyotypes in some subgroups had longer OS with AZA vs CCR: median OS in pts with del 5/5q-, del 5/5q- WITHOUT del 7/7q-, or trisomy 8 as part of a complex karyotype treated with AZA survived 5.1, 8.0, and 12.4 months longer, respectively, than their counterparts who received CCR. HRs with AZA vs CCR in pts with complex cytogenetics ranged from 0.42 (95%CI: 0.10 – 1.69) to 0.55 (95%CI: 0.29 – 1.05). Conclusions These findings support earlier data showing effectiveness of AZA in higher-risk MDS pts with complex or non-complex karyotypes. Major gains in OS were obtained with AZA vs CCR (12-18 months longer OS with AZA) for the following categories: del 7/7q- (non-complex), del 7/7q- WITHOUT del 5/5q- (non-complex), and trisomy 8 (non-complex and complex). Pts with trisomy 8 treated with AZA experienced a 3-fold increase in median OS compared with similar pts who received CCR. Longer OS (AZA 15.3 vs CCR 7.3 months) was also obtained for pts with del5/5q- WITHOUT del7/7q- as part of a complex karyotype. The worse cytogenetic categories, del 7/7q- and del 5/5q- AND del 7/7q-, both with complex karyotype, were associated with the poorest OS regardless of treatment. Pt subgroups in this post hoc analysis were small and heterogeneous; confirmation of these findings in larger pt samples is warranted. Disclosures Mufti: Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Gore:Celgene: Consultancy, Equity Ownership, Research Funding; Johnson & Johnson: Research Funding. Santini:Celgene: Honoraria. Fenaux:Celgene: Honoraria, Research Funding; Ortho Biotech: Honoraria, Research Funding; Roche: Honoraria, Research Funding; Amgen: Honoraria, Research Funding; Cephalon: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; MSD: Honoraria, Research Funding; Epicept: Honoraria, Research Funding. Skikne:Celgene: Employment, Equity Ownership. Hellstrom-Lindberg:Celgene: Research Funding. Seymour:Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Beach:Celgene: Employment, Equity Ownership. Backstrom:Celgene: Employment, Equity Ownership. Fernando:Celgene: Employment, Equity Ownership.

Prognostic Value of “Monosomal Karyotype” in Comparison to “Complex Aberrant Karyotype” in AML: A Study on 824 Cases with Aberrant Karyotype

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 418-418
Author(s):  
Claudia Haferlach ◽  
Tamara Alpermann ◽  
Susanne Schnittger ◽  
Wolfgang Kern ◽  
Torsten Haferlach
Keyword(s):  
Cox Regression ◽  
Complex Karyotype ◽  
Employment Equity ◽  
Cytogenetic Abnormalities ◽  
Cox Regression Analysis ◽  
Poor Risk ◽  
Risk Patients ◽  
Equity Ownership ◽  
The Impact ◽  
Monosomal Karyotype

Abstract Abstract 418 Background: Several classifications based on cytogenetics have been proposed in AML. Typically 3 major categories for prognostication are defined: favorable, intermediate and unfavorable. The assignment to the unfavorable group shows minor differences between the different cytogenetic classifications currently used, however certain cytogenetic subgroups are assigned to the unfavorable subgroup concordantly: −5/5q−, 7q−/−7, −17/abn17p, inv(3)(q21q26)/t(3;3)(q21;q26) and complex karyotype (CK). With respect to CK 3 definitions are used: ≥3, ≥4 or ≥5 unrelated abnormalities. Recently, a so-called “monosomal karyotype” (MSK) defined as a karyotype showing “two or more distinct autosomal chromosome monosomies or one single autosomal monosomy in the presence of structural abnormalities” was introduced (Breems et al. JCO 2008). It was suggested that patients with MSK have a poor outcome being even inferior to CK. Aim: We here evaluated the prognostic power of differently defined cytogenetic subsets in order to identify the best definition for the prognostically most unfavorable subgroup. Patients: From our initial cohort of newly diagnosed AML (n=1,959) patients with t(15;17), t(8;21) or inv(16) (n=170) and AML with normal karyotype (n=965) were excluded. Thus, 824 patients with cytogenetic abnormalities remained for further investigation. Results: 428/824 (51.9%) patients showed an intermediate risk karyotype according to revised MRC criteria (MRC-I) (Grimwade et al. Blood 2010), while the remaining 396/824 (48.1%) cases belonged to the unfavorable MRC group (MRC-U). 162/824 cases (19.7%) fulfilled the criteria of MSK. According to MRC, 4 of these 162 cases with MSK were classified MRC-I while 158 were classified MRC-U. The overlap in classification between CK and MRC-U differed depending on the number of aberrations used to define CK. As such, the number of cases with CK was 272 (33.0%; MRC-I: 17, MRC-U: 255) using ≥3 clonal aberrations, and decreased to 222 (26.9%; all MRC-U) patients using ≥4 clonal aberrations or 196 (23.8%; all MRC-U) cases when applying the criterion of ≥5 clonal aberrations, respectively. Univariate Cox regression analysis revealed that unfavorable cytogenetics as defined by MRC-U, MSK, CK defined as ≥3, ≥4 or ≥5 unrelated abnormalities were all significantly associated with inferior OS as compared to the respective remaining intermediate group (for all p<0.001). Hazard ratios were 1.61, 1.93, 1.68, 1.94, and 1.92, respectively. Median OS in the respective categories was 8.5, 5.7, 6.3, 5.7, and 5.7 months, respectively. We then performed further analyses within the unfavorable risk group defined according to MRC and tested the impact of the 4 definitions for unfavorable subsets. In each comparison the median OS was significantly shorter for the subset with MSK, or CK defined as ≥3, '4 or ≥5 unrelated abnormalities as compared to the remaining MRC-U cases (5.7 vs 11.7 mo p=0.005; 6.3 vs 10.6 mo, p=0.031; 5.7 vs 11.0 mo, p=0.003; 5.7 vs 10.9 mo, p=0.006). Furthermore OS of patients within MRC-U excluding cases with MSK, or CK with ≥3, ≥4 or ≥5 unrelated abnormalities did not differ from patients with cytogenetic abnormalities assigned to MRC-I (median OS 11.7, 10.6, 11.0 and 10.9 mo, respectively vs 21.1 mo, p=0.072, p=0.16, p=0.28, and p=0.11, respectively). Within the MRC-U cohort only 124 cases fulfilled both criteria: MSK and CK≥4 (median OS 5.3 mo), 97 were CK≥4 only (median OS 6.3 mo) and 35 MSK only (median OS 6.7 mo). OS did not differ between these 3 subgroups but was significantly shorter for all comparisons to patients included in none of these subgroups (p<0.001, p=0.009, p=0.012, respectively). On the other hand OS of the 33 cases with 3 unrelated abnormalities did not differ from MRC-U cases with 1 or 2 abnormalities (18.9 vs 10.6, p=0.48). Conclusions: All definitions of very poor risk AML patients allow to identify a subset within MRC-U that shows significantly shorter OS than the remaining MRC-U cases. However, “complex karyotype defined as ≥4 unrelated abnormalities” is the best parameter as it identifies the largest proportion of very poor risk patients. Even more important, the application of the monosomal karyotype for prognostication and clinical guidance in AML misses 24.5% of the very poor risk patients identified based on CK ≥4. This may lead to suboptimal treatment decisions in this clinically proven very high risk patients. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. 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.

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