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

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

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

Blood ◽

10.1182/blood.v116.21.3596.3596 ◽

2010 ◽

Vol 116 (21) ◽

pp. 3596-3596

Author(s):

Claudia Haferlach ◽

Frank Dicker ◽

Susanne Schnittger ◽

Wolfgang Kern ◽

Torsten Haferlach

Keyword(s):

Multivariate Analysis ◽

Regression Analysis ◽

Chronic Lymphocytic Leukemia ◽

Cox Regression ◽

Lymphocytic Leukemia ◽

Employment Equity ◽

Mutation Status ◽

Cox Regression Analysis ◽

Igh Locus ◽

Equity Ownership

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

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In Chronic Lymphocytic Leukemia the Gain of the Short Arm of Chromosome 2 Is Highly Associated with an Unmutated IGHV status, 11q/ATM Deletion, SF3B1 Mutation and a Complex Karyotype

Blood ◽

10.1182/blood.v128.22.4379.4379 ◽

2016 ◽

Vol 128 (22) ◽

pp. 4379-4379

Author(s):

Claudia Haferlach ◽

Sabine Jeromin ◽

Anna Stengel ◽

Manja Meggendorfer ◽

Melanie Zenger ◽

...

Keyword(s):

Multivariate Analysis ◽

Cox Regression ◽

Prognostic Impact ◽

Chromosome 2 ◽

Complex Karyotype ◽

Employment Equity ◽

Cox Regression Analysis ◽

Tp53 Mutations ◽

Equity Ownership

Abstract Background: CLL is characterized by a distinct pattern of translocations, genomic gains and losses and molecular mutations. The most frequent abnormalities such as trisomy 12 and deletions of 6q, 11q, 13q and 17p have been intensively studied. However, data on less frequent recurrent abnormalities such as the partial gain of the short arm of chromosome 2 is lacking. Aims: a) Determine the frequency of 2p gain in CLL, b) Characterize the size and the commonly gained region, c) Analyze the spectrum of additional cytogenetic abnormalities and molecular mutations, and d) Evaluate the prognostic impact. Patients and Methods: Chromosome banding analysis (CBA) revealed a gain of 2p in 113 out of 5564 (2%) CLL cases. In 72 cases with sufficient material genomic array analysis (SurePrint G3 ISCA CGH+SNP Microarray, Agilent, Waldbronn, Germany) and determination of the mutation status of TP53, SF3B1 and IGHV were performed. Results:76% of patients with gain of 2p were male. The median WBC count was 33,700/µL (range: 5,900 - 228,000). Median age was 66 years (range: 29 - 87). The gain of 2p always encompassed the 2p telomere (2pter) while the centromeric border of the 2p gain varied between 2p21 and the centromere of chromosome 2 (2p10) (genomic positions 45,859,076 to 92,297,003). The gain of 2p was the sole chromosomal abnormality in only 8/72 cases (11%) and was accompanied by one, two or more than two additional aberrations in 10, 20, and 34 cases. In total 209 chromosome abnormalities were observed in addition to the 2p gain (median per patient: 2, range: 0-16). Of these only 21 were balanced while 188 were unbalanced abnormalities leading to gain or loss of chromosomal material. Gain of 2p was most frequently accompanied by deletions in 13q (total: 74%, homozygous: 11%), 11q (56%), 18p (18%), and 6q (13%) and gains of 8q (11%). 17p deletions were present in 6% of cases. In 49 cases (68%) the gain of 2p was present in the main clone while it was present in a subclone only in 23 cases (32%). The gain of 2p material was due to a duplication in the short arm of chromosome 2 in 10 cases, while a gain of an isochromosome 2p was present in 3 cases. In the remaining cases material of the short arm of chromosome 2p was attached to a variety of different partner chromosomes. The most frequent acceptor chromosome was chromosome 18 (n=13; 18%). In two cases (2%) 2 IGH rearrangements were observed of which one was mutated and the other unmutated. The IGHV status was unmutated (IGHV-U) in 66 (92%) and mutated in only 4 cases (6%). Three of these 4 cases with mutated IGHV showed only a low mutation rate (sequence homology to germline 97-97.9%). Stereotyped B-cell receptors were present in 14 cases (19%). SF3B1 mutations were observed in 21 cases (29%) with a median mutation load (ML) of 39% (range: 10-51%). TP53 mutations were detected in 8 (11%) cases (median ML: 60%, range: 13-100%). In 2 patients with a TP53 mutation a TP53 deletion was present and in 3 cases a copy neutral loss of heterozygosity (CN-LOH) of 17p was detected leading to TP53 wild-type loss in these 5 cases. TP53 mutations were less frequent in cases harboring the gain of 2p as the sole abnormality (3% vs 21%, p=0.02) The prognostic impact of 2p gain was evaluated in an unselected cohort of 1381 CLL cases with available follow up data (median follow up: 5.1 years) including 22 cases with 2p gain. The frequency of IGHV-Ustatus, SF3B1 mutations and 11q/ATM deletions was significantly higher in CLL with 2p gain compared to cases without (for all p<0.05). In univariate Cox regression analysis gain of 2p was significantly associated with shorter overall survival (OS) (relative risk (RR): 2.1; p=0.05). 5 year OS was 69% in CLL with 2p gain compared to 85% in cases without 2p gain (p=0.05). However, in multivariate analysis only IGHV-U, mutations in SF3B1 and TP53 and TP53/17p deletion were independently associated with shorter OS, while gain of 2p and 11q/ATM deletion were not. 2p gain was associated with shorter time to treatment (TTT) (RR: 2.0; p=0.02). In multivariate analysis only IGHV-U, SF3B1 mutation and 11q/ATM and TP53/17p deletion were independently associated with shorter TTT, while gain of 2p and TP53 mutations were not. Conclusions:CLL with gain of 2p is highly associated with an unmutated IGHV status (92%), a high frequency of 11q/ATM deletion (56%), 13q deletion (74%), SF3B1 mutation (29%) and a complex karyotype (47%). Data suggest that gain of 2p is a later event in CLL pathogenesis and might be a marker of progression. Disclosures Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Jeromin:MLL Munich Leukemia Laboratory: Employment. Stengel:MLL Munich Leukemia Laboratory: Employment. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Zenger:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

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

Blood ◽

10.1182/blood.v118.21.968.968 ◽

2011 ◽

Vol 118 (21) ◽

pp. 968-968 ◽

Cited By ~ 1

Author(s):

Claudia Haferlach ◽

Melanie Zenger ◽

Tamara Alpermann ◽

Susanne Schnittger ◽

Wolfgang Kern ◽

...

Keyword(s):

Overall Survival ◽

Chromosome Banding ◽

Cox Regression ◽

Clonal Evolution ◽

Employment Equity ◽

Cox Regression Analysis ◽

Banding Analysis ◽

Chromosome Banding Analysis ◽

Equity Ownership ◽

Acquired Abnormalities

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

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Higher Expression of Nuclear Cereblon in Bone Marrow Biopsies of Patients with Multiple Myeloma Treated with Imids in the HOVON-87/Nmsg-18 Trial Is Associated with Longer PFS and OS

Blood ◽

10.1182/blood-2019-128584 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 3071-3071

Author(s):

Ruth Wester ◽

M Duin ◽

King Hong Lam ◽

Suzana Couto ◽

Yan Ren ◽

...

Keyword(s):

Board Of Directors ◽

Research Funding ◽

Cox Regression ◽

Employment Equity ◽

Advisory Committees ◽

Cytoplasmic Staining ◽

Cox Regression Analysis ◽

Cytogenetic Aberrations ◽

Equity Ownership ◽

Response Groups

Introduction Response to treatment in patients with multiple myeloma (MM) is variable. With increasing possibilities of treatment regimens, predictive factors for response are important. Immune modulating agents (IMiDs) require Cereblon (CRBN) for activity. Therefore, the aim of this study was to identify the genes involved in the CRBN pathway which predict the response to therapy with IMiDs. Methods Paraffin embedded bone marrow (BM) biopsies were used from newly diagnosed patients included in HOVON-87/NMSG-18 trial obtained at inclusion. In this trial, elderly patients with MM were randomized between treatment with Melphalan-Prednisone (MP)-Thalidomide (MPT) followed by thalidomide maintenance versus MP-Lenalidomide (MPR) followed by lenalidomide maintenance (Zweegman et al. Blood 2016;127:1109-1116). BM biopsies were stained with a fully automated dual color, bright-field immunohistochemical assay for CRBN, its neosubstrates Ikaros and Aiolos and the downstream targets IRF4 and c-MYC. CD138 was used to identify MM plasma cells in the BM samples. For CRBN, both nuclear and cytoplasmic staining was evaluated. The distribution and intensity of the immunostaining was assessed using the H-score. The H-scores were calculated using the following formula: [1 × (% cells 1+) + 2 × (% cells 2+) + 3 × (% cells 3+)] and range from 0-300 (0-600 for combined cytoplasmic-nuclear CRBN H-score). For the Cox regression analysis H-scores were corrected by dividing these by a factor 100: hazard rates were considered per 100 points increase of the H-score. Protein levels of the CRBN pathway were compared between patients with complete response (CR) or very good partial response (VGPR) vs partial response (PR) and no change/progressive disease (NC/PD). High-risk cytogenetic aberrations (FISH) were defined as having deletion of 17p, and/or translocation t(4;14) and/or t(14;16). Statistical analysis was done using univariate and multivariate Cox regression analysis for progression free survival (PFS) and overall survival (OS), and the Mann-Whitney test for comparing response groups. Kaplan-Meier survival curves were generated to illustrate survival. Results BM samples obtained at diagnosis from 149 patients were evaluated. Seventy-one patients were treated in the thalidomide arm vs 78 patients in the lenalidomide arm. Median age was 73 years [range 60-90]. Revised ISS stages I/II/III were 12%/80%/8% respectively. At the time of analysis, median follow up of the 45 patients still alive was 83 months [range 23 - 114 months]. Best response on protocol treatment was sCR/CR in 22%, VGPR in 30%, PR in 36% and NC/PD in 12%. Protein expression across the response groups showed higher nuclear CRBN in patients who responded better (sCR/CR/VGPR; median H-score: 178 (49-273)) compared to patients with a worse response (PR/NC/PD; median H-score: 157 (67-251)), albeit not statistically significant (Mann-Whitney p-value=0.06). Higher H-score of nuclear staining of CRBN was associated with a longer PFS and OS, with a hazard ratio (HR) of 0.52 for PFS (95% confidence interval (CI)=0.37-0.86, p<0.001) and a HR of 0.56 for OS (95% CI=0.36-0.78; p<0.01). Patients with the top quartile nuclear CRBN levels had a median PFS of 21 months longer compared to patients with the lowest quartile (38 months vs 17 months). In terms of OS, patients with the highest quartile nuclear CRBN expression demonstrated a median survival that was 2 times as high as found in patients with the lowest quartile (75 months vs 35 months; Figure 1). In addition, cytoplasmic staining of CRBN was associated with improved PFS (HR = 0.66 (95% CI=0.47-0.94; p=0.02), but not with OS (HR = 0.73 (CI=0.48-1.11); p=0.14). None of the other markers were associated with survival. In a multivariate analysis (which included study arm (MPT vs MPR), nuclear CRBN, high-risk cytogenetic aberrations and R-ISS), nuclear CRBN remained independently associated with OS as well as R-ISS and study arm. For PFS, only nuclear CRBN remained statistically significant after backward selection. Despite treatment arm being a statistically significant term in the multivariate Cox model for OS, no relation was found for treatment arm and nuclear CRBN, in terms of OS. Conclusions In this study we demonstrate that higher expression of nuclear CRBN in myeloma cells in BM of patients with MM was associated with a superior PFS and OS. Disclosures Couto: Celgene Corporation: Employment, Equity Ownership, Patents & Royalties. Ren:Celgene Corporation: Employment, Equity Ownership. Wang:Celgene Corporation: Employment, Equity Ownership. Thakurta:Celgene: Employment, Equity Ownership. Zweegman:Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding. Broyl:Celgene, amgen, Janssen,Takeda: Honoraria. Sonneveld:Amgen: Honoraria, Research Funding; BMS: Honoraria; Celgene: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Karyopharm: Honoraria, Research Funding; Takeda: Honoraria, Research Funding; SkylineDx: Research Funding.

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

Blood ◽

10.1182/blood.v118.21.418.418 ◽

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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Comparison of TP53 Alterations in Hematological Malignancies

Blood ◽

10.1182/blood.v126.23.4819.4819 ◽

2015 ◽

Vol 126 (23) ◽

pp. 4819-4819

Author(s):

Anna Stengel ◽

Wolfgang Kern ◽

Torsten Haferlach ◽

Susanne Schnittger ◽

Melanie Zenger ◽

...

Keyword(s):

Burkitt Lymphoma ◽

Tp53 Mutation ◽

Hematological Malignancies ◽

Complex Karyotype ◽

Mutation Load ◽

Employment Equity ◽

Copy Number State ◽

Tp53 Mutations ◽

Cytogenetic Aberrations ◽

Equity Ownership

Abstract Background: TP53 is altered in ~50% of human cancers. Alterations include mutations and deletions. Both frequently occur together, supporting the classical "two-hit" hypothesis for tumor-suppressor genes. Aim: Comparison of TP53 mutation/deletion patterns in different hematological malignancies, including AML, MDS, ALL, Burkitt lymphoma, CLL and T-PLL. We analyzed (i) the frequencies of TP53 mutations and deletions, (ii) the types of mutation, (iii) the mutation load, (iv) the correlations to cytogenetic aberrations, (v) the age dependency, and (vi) impact on survival. Patient cohort and methods: A total of 3383 cases (AML: n=858, MDS: n=943, ALL: n=358, Burkitt lymphoma: n=25, CLL: n=1148 and T-PLL: n=51) were analyzed for TP53 deletions by interphase FISH determining the copy number state and for TP53 mutations by next-generation amplicon deep sequencing. Karyotype data was available for all cases. Results: Overall, alterations in TP53 were detected in 361/3383 cases (11%; 186 cases with mutation only (mut only), 51 cases with deletion only (del only), 124 cases with mutation and deletion (mut+del)). Regarding the respective entities, the highest frequency of TP53 alterations was observed in patients with Burkitt lymphoma (total alteration frequency: 56%, mut+del: 12%, mut only: 44%, no case del only). Alterations in TP53 also occured with a high incidence in patients with T-PLL (total: 30%; mut+del: 10%; mut only: 4%; del only: 16%) followed by cases with ALL (total: 19%; mut+del: 6%; mut only: 8%; del only: 5%) and AML (total: 13%; mut+del: 5%; mut only: 7%; del only: 1%). By contrast, TP53 alterations occurred less frequently in patients with CLL (total: 8%; mut+del: 4%; mut only: 3%; del only: 1%) and MDS (total: 7%; mut+del: 1%; mut only: 5%; del only: 1%). Missense mutations were found to be the most abundant mutation type in all entities analyzed with a frequency ranging from 71% - 88%. In all entities mainly one mutation per case was detected; however, MDS cases were found to harbour a statistically increased proportion of cases with two mutations compared to the other entities (p = 0.003). High TP53 mutation loads were detected in T-PLL (median: 88%) and AML (47%), whereas the lower ones were found in ALL (28%), Burkitt lymphoma (39%), MDS (39%), and CLL (36%). A strong correlation of TP53 alterations with a complex karyotype was observed in AML (of patients with TP53 alteration: 5% with normal karyotype, 67% with complex karyotype, 28% with other aberrations), ALL (16% normal, 45% complex, 39% other), MDS (14% normal, 53% complex, 33% other), and T-PLL (20% normal, 47% complex, 33% other). By contrast, in CLL and Burkitt lymphoma, TP53 alterations were mainly correlated with other aberrations (CLL: 10% normal, 30% complex, 60% other; Burkitt: 29% normal, 0% complex, 71% other). TP53 mut and TP53 mut+del were significantly more frequent in patients ≥ 60 vs < 60 years in AML (9% vs. 2% for mut only, p < 0.001; 7% vs. 2% for mut+del, p = 0.001) and ALL (12% vs. 6% for mut only, p < 0.001; 13% vs. 3% for mut+del, p = 0.001). By contrast, no such differences were observed for patients with CLL, MDS, T-PLL and Burkitt lymphoma. Moreover, TP53 alterations (especially of TP53 mut+del) had a significant negative impact on OS in all entities except for T-PLL and Burkitt lymphoma, most probably due to their overall short OS or due the lower number of cases. Conclusion: The frequency of TP53 mutations and/or deletions as well as the mutation load clearly varied between different hematological malignancies with the highest incidence of TP53 mut in patients with Burkitt lymphoma (56%) and a rather low frequency in CLL (7%) and MDS (6%). TP53 del were frequent in patients with T-PLL (26%) and Burkitt lymphoma (12%) and are hardly found in MDS cases (2%). TP53 alterations are correlated to higher age in AML and ALL. Moreover, alterations in TP53 are correlated to a short OS and to a complex karyotype, with the exception of Burkitt lymphoma and CLL, were they were found to be associated to other cytogenetic aberrations. Thus, TP53 mutations and deletions need further investigation in the future, especially regarding their clinical impact in different hematologic entities. Disclosures Stengel: 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. Zenger:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

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A Comprehensive Panel of Molecular Mutations Notably Improves a Cytogenetic Prognostication System in Routine AML Diagnostics

Blood ◽

10.1182/blood.v128.22.286.286 ◽

2016 ◽

Vol 128 (22) ◽

pp. 286-286

Author(s):

Claudia Haferlach ◽

Manja Meggendorfer ◽

Annette Fasan ◽

Karolina Perglerová ◽

Wolfgang Kern ◽

...

Keyword(s):

Molecular Genetics ◽

Risk Group ◽

Classification Systems ◽

Prognostic Impact ◽

Intermediate Risk ◽

Complex Karyotype ◽

Employment Equity ◽

Mutation Status ◽

Cox Regression Analysis ◽

Equity Ownership

Abstract Background: Based on sequencing studies the molecular landscape of AML has been unraveled. Novel prognostic scores combining molecular mutations and karyotype have been proposed (Grimwade et al. Blood, 2016, Döhner et al. NEJM 2015). However, these proposed classification systems differ in several aspects and yet no consensus has been established which genetic information is required for prognostication in AML today. Aims: 1) Test the prognostic value of a panel of molecular markers in addition to a cytogenetic score in a large cohort of AML patients. 2) Determine the proportion of patients with a suitable molecular marker for disease monitoring (MRD) applying molecular genetics. Patients and Methods: 867 de novo AML cases younger than 60 years were investigated (median age: 48 years, median follow up of 41 months). All patients were evaluated for karyotype, KMT2A-PTD, FLT3-ITD and in addition for mutation status of ASXL1, CEBPA, DNMT3A, NPM1, RUNX1 and TP53 according to the proposal by Grimwade et al. Blood 2016. Results: First, AML were classified according to the refined MRC cytogenetic classification with AML with t(15;17)/PML-RARA regarded as a separate group (n=89 (10%), 90% overall survival (OS) at 5 years). 89 cases (10%) were assigned to the favourable risk group (t(8;21)/RUNX1-RUNX1T1: n=42; inv(16)/t(16;16)/CBFB-MYH11: n=47), 570 (68%) to the intermediate risk group and 119 (14%) to the adverse risk group. OS at 5 years was 66%, 53% and 28%, respectively, and differed significantly between all four subgroups (for all comparisons p<0.001). Next, the following subgroups were separated: CEBPA double mutated (dm) cases (n=44 (5%); OS at 5 years: 83%), NPM1mut/FLT3-ITD- AML (n=181 (21%); OS at 5 years: 62%), and NPM1mut/FLT3-ITD+ AML (n=137 (16%); OS at 5 years: 47%; for all comparisons between these 3 groups p<0.001). Thus, prognosis of CEBPAdm cases was comparable to PML-RARA+ AML. OS in NPM1mut/FLT3-ITD- AML is comparable to CBF-leukemias. In NPM1mut AML no prognostic impact of DNMT3Amut was found. In all these 3 groups defined on molecular genetics no prognostic impact of additional karyotype information on OS was observed. Next, in the remaining cases of the cytogenetic intermediate risk group (n=209) the prognostic impact of mutations in ASXL1, DNMT3A, RUNX1, TP53, KMT2A-PTD and FLT3-ITD was evaluated. In multivariate Cox regression analysis mutations in TP53 (relative risk (RR): 3.5; p=0.04), ASXL1 (RR: 2.2, p=0.004), and FLT3-ITD (RR: 1.8; p=0.04) were independently associated with shorter OS. OS at 5 years was 25% in cases carrying at least one of these mutations compared to 54% in cases with none of these mutations (p=0.001). Within the adverse cytogenetic risk group cases with either a complex karyotype (n=27) or a KMT2A (n=25) or MECOM rearrangement (n=14) had the worst outcome compared to the remaining cases (OS at 5 yrs: 19% vs 54%, p=0.02). In the remaining cases the presence of at least one mutation in either ASXL1, TP53 or FLT3-ITD was associated with worse outcome (OS at 5 yrs: 33% vs 74%, p=0.04). Thus, AML with complex karyotype, KMT2A or MECOM rearrangements had the worst prognosis, while cases with adverse cytogenetics and at least one mutation in either ASXL1, TP53 or FLT3-ITD have a slightly better outcome which is comparable to AML with intermediate risk cytogenetics harbouring one of these mutations. OS in AML with adverse cytogenetics without mutations in ASXL1, TP53 or FLT3-ITD is not worse than in AML with intermediate cytogenetics also lacking these mutations. A fusion gene or a molecular mutation as target for MRD monitoring was present in 791 patients (91%) when all genes analysed were taken into account. If only markers showing prognostic relevance, i.e. fusion genes, mutations in NPM1, CEBPA, FLT3-ITD, ASXL1 and TP53 were considered a MRD marker was still available in 726 cases (84%). Conclusions: 1) In AML a prognostication system is feasible based on the identification of t(15;17)/PML-RARA, t(8;21)/RUNX1-RUNX1T1, inv(16)/t(16;16)/CBFB-MYH11, 11q23/KMT2A rearrangements, 3q26/MECOM rearrangements, complex karyotype, and mutation status of NPM1, CEBPA, ASXL1, TP53, and FLT3-ITD (figure 1). 2) The analysis of these parameters allows to identify an MRD marker in 84% of patients. 3) The analysis of additional genes may be required in a comprehensive AML work-up as soon as novel targeted treatment strategies will become available. Disclosures Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Fasan:MLL Munich Leukemia Laboratory: Employment. Perglerová:MLL2 s.r.o.: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

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Higher Percentages of Ring Sideroblasts and SF3B1 Mutations in Patients with AML Correlate with Mutations in TP53 and Adverse Cytogenetics, but Have No Independent Impact on Outcome

Blood ◽

10.1182/blood.v126.23.3847.3847 ◽

2015 ◽

Vol 126 (23) ◽

pp. 3847-3847

Author(s):

Pedro Martin-Cabrera ◽

Sabine Jeromin ◽

Tamara Alpermann ◽

Karolína Perglerová ◽

Claudia Haferlach ◽

...

Keyword(s):

Bone Marrow ◽

De Novo ◽

Wild Type ◽

Employment Equity ◽

Tp53 Mutations ◽

Ring Sideroblasts ◽

Impact On Survival ◽

Equity Ownership ◽

Kras Codon ◽

Codon 61

Abstract Introduction Ring sideroblasts (RS) are common findings in myeloid malignancies as MDS or MDS/MPN, especially in RARS-T, and are associated with SF3B1 mutations. However, the incidence of both RS and SF3B1 mutations have not yet been assessed properly in acute myeloid leukemia (AML). Aim Determine the frequency of RS and SF3B1 mutations in 1857 patients with de novo and therapy-related AML (t-AML). Define their impact on survival and association with other frequently mutated genes, as well as with cytogenetic abnormalities. Patients and Methods From a total of 1857 AML patients (excluding those with cytogenetically-defined entities according to WHO), bone marrow assessment revealed 473 (25%) with RS ≥ 1% and thereof 183 (10% of all) with ≥15 RS. Sequencing or melting curve analysis were performed in a subcohort of 340/473 patients for the detection of mutations in: SF3B1, ASXL1, DNMT3A, FLT3- TKD, IDH1 R132, IDH2 R140, IDH2 R172, KRAS, NPM1, NRAS, RUNX1, TET2, FLT3 -ITD and MLL -PTD. These 340 cases were subject to the study. Out of these 340, 141 cases (42%) had RS ≥ 15% and the remaining 199/340 (59%) had RS ≥1 to <15%. The cohort consisted of 303 (89%) de novo AML (FAB: M0 n=18, M1 n=67, M2 n=165, M4 n=30, M5 n=3, M6 n=20) and 37 (11%) t-AML. 148 were female and 192 were male, with median age 74 years, range: 20-93 years. Chromosome banding analysis (assisted by FISH if needed) was performed in all 340 cases. Results The percentage of bone marrow blasts correlated inversely with the percentage of RS present (r 0.213, p<0.001). 136 (40%) patients had a normal karyotype. Intermediate cytogenetics according to MRC criteria were found in 193 (57%), and adverse in 147 (43%). Patients with RS ≥15% more frequently had adverse cytogenetics in comparison to those with RS between 1-14% (54% vs 36%, p=0.001). The frequencies of gene mutations were as follows: TP53 103/331 (31%), RUNX1 84/315 (26%), DNMT3A 86/337 (25%), TET2 68/330 (20%), ASXL1 58/334 (17%), IDH2 R140 53/338 (15%), NPM1 43/340 (12%), SF3B1 34/334 (10%), FLT3 -ITD 33/340 (10%), NRAS 29/340 (8%), IDH1 R132 21/339 (6%), MLL -PTD 22/337 (6%), FLT3 -TKD 18/333 (5%), KRAS codon 12 13/299 (4%), IDH2 R172 13/338 (3%) and KRAS codon 61 3/299 (1%). Moreover, in total 30 variants in 28 patients were identified in DNMT3A, RUNX1, TET2 and TP53, which according to current knowledge cannot be assigned to mutations or SNPs yet. Patients with ≥15% more frequently had TP53 mutations (mut) (44% vs 22%, p<0.001) and less frequently IDH2 R140 mut (11% vs 19%, p=0.094) and MLL -PTD (2% vs 6%, p=0.006). Accordingly, patients with TP53 mutations had higher percentages of RS as compared to those without (28% vs 16%, p<0.001) and patients with IDH2 R140 mut and MLL -PTD, respectively, had lower percentages of RS as compared to those without (15% vs 21%, p=0.043 and 11% vs 21%, p=0.025, respectively). Furthermore, patients with mutations in the following genes had fewer RS than patients with the respective wild-types: ASXL1 (15% vs 21%, p=0.040), FLT3 -ITD (14% vs 21%, p= 0.049), IDH2 R140 (15% vs 21%, p=0.043), MLL -PTD (11% vs 21%, p=0.025), NPM1 (13% vs 21%, p=0.018) and KRAS codon 61 (3% vs 20%, p<0.001). Conversely, patients with mutated SF3B1 had more RS than patients with wild type (27% vs 19%, p=0.054). However, the number of RS did not translate into an increase in the mutational burden of SF3B1. Given the limited degree of overlap between mutations of the four most frequently mutated genes, we hierarchically separated the patients into 5 groups: TP53 mut, ASXL1 mut, NPM1 mut, SF3B1 mut and patients without any of these mutations. Interestingly, the percentage of RS was very similar in the two groups, TP53 mut and SF3B1 mut, and significantly higher as compared to all other groups (TP53 mut/SF3B 1mut: 28% vs ASXL1 mut/NPM1 mut/others 14%, p<0.001). The number of RS did not have an impact on the overall survival (OS) and event free survival (EFS) of patients. Conclusion 1. Ring sideroblasts ≥ 15% are present in 10% of de novo and t-AML. 2. The blast count correlates inversely with the number of RS. 3. Patients with ≥ 15% RS more frequently carry TP53 mutations and adverse cytogenetics. 4. Although patients with ≥ 15% RS have worse molecular (TP53 mut) and cytogenetic features, there is no statistically significant impact on survival when compared to patients with <15% RS. 5. Patients with mutated ASXL1, FLT3 -ITD, IDH2 R140, MLL -PTD and NPM1 have less RS than wild type patients while those with TP53 or SF3B1 mutations have higher RS. Figure 1. Figure 1. Disclosures Martin-Cabrera: MLL Munich Leukemia Laboratory: Employment. Jeromin:MLL Munich Leukemia Laboratory: Employment. Alpermann:MLL Munich Leukemia Laboratory: Employment. Perglerová:MLL2 s.r.o.: 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.

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TP53 Mutations Are the Most Unfavorable Genetic Alteration in AML

Blood ◽

10.1182/blood.v120.21.660.660 ◽

2012 ◽

Vol 120 (21) ◽

pp. 660-660 ◽

Cited By ~ 1

Author(s):

Claudia Haferlach ◽

Vera Grossmann ◽

Christiane Eder ◽

Alexander Kohlmann ◽

Wolfgang Kern ◽

...

Keyword(s):

Cox Regression ◽

Close Association ◽

Tp53 Gene ◽

Dominant Negative ◽

Dominant Negative Effect ◽

Complex Karyotype ◽

Cox Regression Analysis ◽

Tp53 Mutations ◽

Equity Ownership ◽

The Impact

Abstract Abstract 660 Background: TP53 is the most frequently mutated gene in cancer. An association of TP53 mutations and adverse prognosis was shown in multiple malignancies. In AML, a high frequency of TP53 alterations has been reported in cases with complex karyotype. However, thus far, a comprehensive study analyzing the impact of TP53 alterations has been lacking. Aims: 1. Determine the frequency of TP53 mutations (TP53mut) and TP53 deletions (TP53del) in a large cohort of AML. 2. Analyze the relation of TP53mut and TP53del with cytogenetics and other molecular mutations. 3. Evaluate the impact of TP53 alterations on outcome. Patients and methods: In 1,000 AML patients (median age 66.8 yrs) the TP53 gene (exons 4–11) was analyzed to detect mutations by either DHPLC with subsequent direct Sanger sequencing (n=190) or a next-generation amplicon deep-sequencing assay (n=810) (454 Life Sciences, Branford, CT). All cases with available material (n=858) were analyzed by interphase FISH for TP53del. In all cases the karyotype was available and categorized according to the refined MRC classification (Grimwade et al., Blood 2010). Cases were also screened for mutations in NPM1 (n=966), CEPBA (n=997), RUNX1 (n=907), ASXL1 (n=937) as well as for FLT3-ITD (n=999) and MLL-PTD (n=952). Clinical follow-up data was available in 841 patients. Results: Frequency of TP53 mutations and deletions: In 115 patients (11.5%) a total of 131 TP53mut were detected. 99 patients showed one and 16 cases two TP53 mutations. Heterozygous deletions of the TP53 gene were detected by FISH in 55/858 (6.4%) patients. In 97/115 cases with TP53mut also the TP53del status was available: 41/97 (42.3%) cases harbored both a TP53mut and a TP53del. 32 of the 56 (57.1%) TP53mut cases without TP53del showed heterozygous and 24 (42.8%) homozygous TP53mut. 13/32 (40.6%) cases with heterozygous mutations harbored two distinct TP53mut, whereas only 19/32 (59.3%) were affected by one mutation suggesting a dominant negative effect of these mutations. In patients with homozygous mutations and no TP53del a copy neutral loss of heterozygosity (CN-LOH) can be assumed. In 2 of these patients SNP microarray data was available revealing in both cases a CN-LOH spanning from 17p11.2 to 17p13.3. Association with cytogenetics and other molecular markers: TP53mut were observed in 1/106 cases with favorable, 12/688 with intermediate (1.7%), and 17/90 (18.9%) with adverse cytogenetics. In cases with complex karyotype TP53mut frequency was 73.3% (85/116). TP53mut were mutually exclusive of CEPBA and NPM1 mutations. In patients harboring TP53mut FLT3-ITD, MLL-PTD, RUNX1 and ASXL1 mutations were detected at low frequencies (2.6%, 4.3%, 7.8% and 4.3%, respectively). TP53del were observed in 2/88 (2.3%) patients with favorable, in 6/601 (1.0%) with intermediate, and in 47/169 (27.8%) cases with adverse cytogenetics. 39/47 (83.0%) patients with adverse cytogenetics and TP53del harbored a complex karyotype. 41/55 (74.5%) cases with TP53del also harbored a TP53mut. Clinical impact: Median OS in patients with TP53mut (n=80) vs TP53 wild-type (wt) cases (n=761) was 4.6 vs 35.6 months (mo) (P<0.001), median EFS was 3.1 vs 13.3 mo (P<0.001); OS and EFS at 3 yrs was 0% vs 49.6% and 0% vs 33.8%. Within the complex karyotype cohort, patients with TP53mut (n=56/80) showed an inferior outcome compared to TP53wt cases (n=24/80) (OS and EFS at 3 yrs: 0% vs 27.9%, P=0.002, 0% vs 25.7%, P=0.002). In univariable Cox regression analyses TP53mut and TP53del were significantly associated with shorter OS (hazard ratio (HR) 3.49; P<0.001 and 2.33; P<0.001). Additionally, in multivariable Cox regression analysis the following parameters were included (which were also significantly associated with OS in univariable analysis): favorable and adverse cytogenetics, complex karyotype, monosomal karyotype, CEPBA double-mutations, NPM1mut/FLT3-ITD-, MLL-PTD, RUNX1, and ASXL1 mutations. Independent prognostic factors were: complex karyotype (HR 1.64, P=0.05), CEPBA double-mutations (HR 0.30; P=0.002), NPM1mut/FLT3-ITD- status (HR 0.62; P=0.004), ASXL1mut (HR 1.46; P=0.016), and TP53mut (HR 2.17; P<0.001). Conclusions: 1. TP53mut occurred in 11.5% and TP53del in 6.4% of AML patients and both showed a close association with adverse karyotype, especially complex karyotype. 2. TP53mut was the parameter with highest risk for adverse outcome compared to any other known genetic risk marker in AML thus far. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Equity Ownership. Grossmann:MLL Munich Leukemia Laboratory: Employment. Eder:MLL Munich Leukemia Laboratory: Employment. Kohlmann:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Equity Ownership.

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TP53 Mutations Are Present In 15.7% Of ALL, Are Especially Frequent In ALL With MYC-Rearrangement and In ALL With Low Hypodiploid Chromosome Status and Are Associated With Poor Prognosis

Blood ◽

10.1182/blood.v122.21.827.827 ◽

2013 ◽

Vol 122 (21) ◽

pp. 827-827

Author(s):

Claudia Haferlach ◽

Sandra Weissmann ◽

Sabrina Kuznia ◽

Susanne Schnittger ◽

Wolfgang Kern ◽

...

Keyword(s):

Mutation Frequency ◽

Tp53 Mutation ◽

Normal Karyotype ◽

Prognostic Impact ◽

Complex Karyotype ◽

Mutation Load ◽

Employment Equity ◽

Tp53 Mutations ◽

Equity Ownership ◽

B Lineage

Abstract Background TP53 is the most frequently mutated gene in cancer. The prognostic impact of TP53 mutations has been demonstrated in CLL, AML, and MDS. However, data on the frequency and prognostic impact of TP53 mutations in ALL is scarce. Aims We aimed at determining the TP53 mutation frequency, the association with cytogenetic subgroups and age, as well as the impact on survival. Patients and Methods In total, a large cohort of 625 patients with ALL was analyzed for TP53 mutations by deep-sequencing allowing to simultaneously quantify the mutation load. In all patients chromosome banding analyses have been performed. In addition, in 341 patients the copy number state of TP53 was determined by FISH. Results The cohort comprised 353 male and 272 female patients, median age was 49.5 years (range: 0.1-91.4 years). The cohort included the following groups: normal karyotype (n=101; 16.2%), t(9;22)(q34;q11) (n=162; 25.9%), MLL-translocations (n=37; 5.9%), MYC-translocations (n=40; 6.4%), t(12;21)(p13;q22) (n=15; 2.4%), low hypodiploidy (<40 chromosomes) (n=24; 3.8%), high hyperdiploidy (51-68 chromosomes) (n=38; 6.1%), complex karyotype (n=69; 11.0%), other cytogenetic abnormalities (n=139; 22.2%). Detailed data on immunophenotyping was available for 408 patients (T-lineage: n=105; B-lineage: n=267, Burkitt: n=36). In the total cohort, the frequency of TP53 mutations was 15.7% (98/625). TP53 mutations were most frequent in ALL with low hypodiploidy (22/24; 91.7%) and MYC-translocated ALL (25/40; 62.5%) and also quite frequent in ALL with complex karyotype (16/69; 23.2%), ALL with normal karyotype (13/101; 17.4%), and in MLL-translocated ALL (6/37; 16.2%). TP53 mutations were rare in t(9;22)(q34;q11) (7/162; 4.3%), high hyperdiploidy (3/38; 6.1%), and other cytogenetic abnormalities (6/139; 4.3%) and absent in ALL with t(12;21)(p13;q22) (0/15; 0%). Furthermore, TP53 mutations were less frequent in T-lineage ALL (8/105; 7.6%) as compared to B-lineage and Burkitt ALL (41/267; 15.4% and 21/36; 58.3%). TP53 mutation frequency increased with age (TP53mut <60 years vs ≥ 60 years 10.8% (45/417) vs 25.5% (53/208), p<0.0001). 25/64 (39.1%) TP53 mutated patients with available TP53 deletion status showed a deletion of the second allele, while in 17/319 (5.3%) TP53wt patients with available TP53 deletion status a TP53 deletion was detected (p<0.001). 11/98 (11.2%) TP53 mutated patients showed two TP53 mutations. Median overall survival (OS) was significantly shorter in TP53mut vs TP53wt patients (18.8 months vs 75.5 months, p<0.0001). OS at 4 years in patients <60 years was 80.1% in TP53wt compared to 56.8% in TP53mut (p=0.012) and 59.3% vs 22.6% in patients ≥ 60 years (p<0.0001). Also within the cytogenetic categories MYC-translocated and complex karyotype TP53 mutations had a significant adverse impact on overall survival. Further, patients with either two TP53 mutations or one TP53 mutation and an accompanying TP53 deletion had a significantly shorter OS as compared to patients with only one altered TP53 allele (median OS 11.5 vs 63.1 months, p=0.009). In contrast, OS in patients with a TP53 deletion without a TP53 mutation did not differ from patients without TP53 alterations. In addition, the TP53 mutation load was investigated by next-generation sequencing and varied between 2% and 98% (median: 41%). Within the subset of patients with TP53 mutation, patients with a mutation load >20% showed a significantly shorter OS as compared to patients with a lower mutation load (median OS 11.5 months vs not reached, p=0.003). Interestingly, OS in patients with a TP53 mutation load ≤ 20% did not differ from TP53wt patients. In multivariate Cox regression analysis including parameters significantly associated with shorter OS in univariate analysis the following factors retained an independent adverse impact on OS: age (<60 years vs ≥ 60 years, HR=2.2; p=0.01), MLL-translocation (HR=2.8; p=0.03), and TP53mut >20% (HR: 3.1, p=0.01). Conclusions 1. TP53 is mutated in 15.7% of ALL with the highest frequency in ALL with low hypodiploidy (91.7%) and MYC-translocated ALL (62.5%). 2. The TP53 mutation frequency increases with age. 3. TP53 mutations are associated with short survival independent of age and specific cytogenetic alterations. 4. TP53 mutations had a significant impact on OS only if the mutation load was >20%. Disclosures: Haferlach: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Weissmann:MLL Munich Leukemia Laboratory: Employment. Kuznia:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kohlmann:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

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