Sole Cytogenetic Abnormality Research Articles

Case report: Pediatric acute lymphoblastic leukemia with trisomy 5 as sole cytogenetic abnormality

Acute lymphoblastic leukemia (ALL) is the most common malignancy in childhood and represents about 75 -80% of ALL in pediatric age group. ALL is characterised by uncontrolled proliferation of abnormal, immature lymphocytes and their progenitors and replacing the bone marrow elements and other lymphoid organs by leukemic cells (ALL blasts). The 5 year survival rate for childhood ALL is about 90% overall, children in lower risk groups have a better prognosis than those in higher risk group. Risk assessment is mainly based on cytogenetic and molecular factors in addition, clinical symptoms and signs, White blood cell count at diagnosis are all recognized for stratification. In present case the cytogenetic analysis showed the presence of trisomy 5 as a sole numerical abnormality. Trisomy 5 accounts for aneuploidy change in the cytogenetic analysis. The gain or loss of whole chromosome, ie aneuploidy is a major genomic insult in human cancers. Aneuploidy is observed in ~90% of solid tumors and~60% of hematological maignancies. The increased gene expression in trisomy 5 causes chromosomal instability (CIN), microsatellite instability (MIN) and genomic instabilty which inturn causes the cancer genome to undergo evolution, adaptation and favors tumor progression in patients with B cell acute lymphoblastic leukemia.

The Molecular Landscape of Acute Myeloid Leukemia with Trisomy 8: A Comprehensive Cancer Center Analysis of 627 Patients

1. Introduction Trisomy 8 is one of the most frequent cytogenetic abnormalities in acute myeloid leukemia (AML). Trisomy 8 frequently occurs with additional aberrant or complex karyotypes; it can also occur as the sole cytogenetic abnormality. While prior studies have shown that trisomy 8 cooperates with other somatic genetic mutations (e.g., RUNX1), there is persistent controversy about the prognostic impact of AML with trisomy 8. We analyzed the relationship between trisomy 8 and additional molecular lesions, the ELN 2022 revision, and the prognostic impact of trisomy 8 across modern treatment strategies. 2. Methods We analyzed 627 patients from the Project ERIS database with newly diagnosed AML treated from January 1, 2013 to April 18, 2023 at VCU Massey Comprehensive Cancer Center. We identified 86 patients with trisomy 8 independent of the presence of complex cytogenetics and a cohort of 541 historical controls without trisomy 8. We recorded baseline patient-related and disease characteristics, including age, ECOG, Charlson comorbidity index (CCI) scores, molecular profiling and ELN 2022 cytogenetic risk, dates of regimen initiation, and survival. We used the D'Agostino & Pearson method for normality testing and the t-test or Mann-Whitney (as applicable) tests for between-group comparisons. Categorical comparisons used Fischer's exact test. We applied the Bonferroni correction if multiple comparisons were made. The Wilson-Brown method was used for 95% confidence intervals. We analyzed survival by the Kaplan-Meier method with significance determined by the log-rank test. The event for calculating the overall survival (OS) was the date of death. Patients were otherwise censored at the date of last contact. 3. Results The most common cytogenetic abnormalities associated with trisomy 8 were the presence of a complex karyotype (45.2%, 95% CI: 35.0-55.9), loss of 5 (15.5%, 95% CI, 9.3-24.7%), loss of 7 (8.3%, 95% CI, 4.1-16.2), and -17p (13.1%, 95% CI, 7.5-21.9). ASXL1 mutations tended to occur significantly more frequently in trisomy 8 compared to those without (27.5% vs 13.0%, p = 0.029, Table A). Similarly, TP53 mutations occurred significantly more frequently in trisomy 8 than those without (27.5% vs 13.0%, p = 0.029). Conversely, DNMT3A mutations were less likely to be associated with trisomy 8, which approached statistical significance (10.0% vs 23.47%, p = 0.063). NPM1 mutations were significantly less likely to be associated with trisomy 8 compared to those without (7.5% vs 30.3%, p = 0.002). Consequently, trisomy 8 was significantly more likely to be associated with ELN 2022 adverse risk disease (odds ratio: 2.754, p < 0.0001). Next, we compared response and survival with intensive chemotherapy using conventional 7+3 in those with and without trisomy 8. In 51 patients with trisomy 8, the composite complete response rate (CRc; CR + CRi + CRh) was 34.1% (95% CI, 21.9-48.9), significantly worse compared to 330 patients without trisomy 8 (55.1%, 95% CI, 49.4-60.6, p = 0.010). The median overall survival was significantly worse in patients with trisomy 8 treated with 7+3 compared to those without trisomy 8 (7.1 m. vs 19.2 m., p = 0.009, Figure B). In 13 patients treated with venetoclax and a hypomethylating agent (HMA), the CRc was 25.0% (95% CI, 8.9-53.2), which was nonsignificantly worse compared with 87 patients without trisomy 8 that demonstrated a CRc of 48.4% (95% CI, 36.4-60.6, p = 0.206). The overall survival of patients treated with venetoclax + HMA nonsignificantly favored patients without trisomy 8 (8.2 m. vs 5.0 m., p = 0.268). 4. Discussion AML with trisomy 8 is significantly enriched in ASXL1 and TP53 mutations; conversely, trisomy 8 is significantly less likely to harbor NPM1 mutations. Patients with trisomy 8 are significantly more likely to have ELN 2022 adverse-risk disease. The CRc rate and median overall survival are significantly worse in patients with trisomy 8 treated with intensive chemotherapy. We observed no significant difference in response or survival in patients with or without trisomy 8 treated with venetoclax + HMA.

Characterization of Cases with the Rare Cytogenetic Abnormality I(7)(p10) Reveals an Association with IDH2 Mutated Acute Myeloid Leukemia

Background: The chromosomal alteration i(7)(p10) is a rare abnormality in hematological malignancies. It leads to a loss of the long arm (7q) and duplication of the short arm (7p) of chromosome 7. However, the presence, frequency and clinical implications of i(7)(p10) in hematological neoplasms has not been analyzed systematically to date. Aim: (1) Analysis of the frequencyand occurrence of i(7)(p10) in hematological malignancies. (2) Characterization of additional cytogenetic alterations and co-occurring mutations. (3) Analysis of potential clinical implications. Patients and methods: All samples sent for diagnosis of suspected hematologic neoplasms between 08/2005 - 05/2023 were used for detection of i(7)(p10). Presence of i(7)(p10) was analyzed using chromosome banding analysis (CBA). In cases with available molecular genetic analyses the following genes were analyzed: ASXL1, BCOR, CEBPA, DNMT3A, EZH2, FLT3-ITD, IDH1, IDH2, KRAS, NPM1, NRAS, RUNX1, SF3B1, SRSF2, STAG2, TET2, TP53, U2AF1 and ZRSR2. Variants of unknown significance were excluded from statistical analysis. All reported p-values are two-sided and were considered significant at p<0.05. Results: In 34 cases an i(7)(p10) was detected. Regarding specific entities, i(7)(p10) was mainly found in AML (23/34, 68%) and rarely in other entities (mature B-cell neoplasms (B-NHL): 6/34, 18%; MDS: 4/34, 12%; ALL: 1/34, 3%). In 20/34 cases, i(7)(p10) comprised the sole cytogenetic abnormality, including 19 AML cases and 1 MDS case. In 10/34 cases, additional chromosomal alterations were detected, leading to a complex karyotype in 5 of these cases. Of note, all cases with a complex karyotype were B-NHL. Additionally, in 4 cases i(7)(p10) was present only in a subclone (1 AML, 1 B-NHL, 2 MDS cases) (Figure A). In 18/34 cases, molecular genetic data was available, revealing DNMT3A (15/18), IDH2 (14/18) and BCOR (7/18) as most frequently mutated genes. As these 18 cases comprised n=15 cases diagnosed as AML, further analysis focused on these patients only. 14/15 cases harbored a mutation in IDH2, the remaining case showed an IDH1 mutation. Further, in 14/15, 7/15 and 4/15 cases, mutations in DNMT3A, BCOR and EZH2 were detected, respectively. By contrast, mutations that are known to frequently co-occur with IDH2 mutations in AML ( SRSF2, ASXL1, NPM1, STAG2, RUNX1, NRAS) were absent or only very rarely detected (Figure B). 12/14 cases with IDH2 mutations showed the mutation site p.Arg172Lys, 2/14 cases p.Arg140Leu, which is known to be the most frequent IDH2 mutation site. This is corroborated by analysis of a control cohort of n=60 IDH2 mutated AML cases without i(7)(p10), in which only 8/60 cases with p.Arg172Lys were detected (86% vs. 13%, p<0.001). Further, comparison of variant allele frequency (VAF) of IDH2 with the VAF of co-mutations suggested the presence of IDH2 in the main clone of all cases, although IDH2 VAF of cases with i(7)(p10) was slightly but not significantly lower than the IDH2 VAF of AML cases without i(7)(p10) (38.3 vs. 45.9). Moreover, comparison of IDH2 mutated AML with and without i(7)(p10) revealed a predominance of females (13/14, 93% vs. 29/60, 48%, p<0.001) and of younger age (63 vs. 72 years, p=0.015) in cases with i(7)(p10). Analysis of overall survival (OS) did not reveal a significant difference between IDH2 mutated AML cases with and without i(7)(p10), although cases with i(7)(p10) showed a trend towards a better outcome (37 vs. 14 months). Conclusions: (1) The rare but recurrent cytogenetic aberration i(7)(p10) is mainly found in AML, where associations with mutations in IDH2, DNMT3A and BCOR were observed. By contrast, other AML-defining mutations ( NPM1, FLT3-ITD, CEPBA) were largely absent, further no defining fusions/rearrangements or a complex karyotype were detected. (2) The vast majority of patients were female and younger than patients with IDH2 mutation without i(7)(p10). (3) Almost all cases showed the rare hotspot p.Arg172Lys and i(7)(p10) as sole aberration, thus it can be speculated that these cases define a distinct subgroup. (4) OS seems to be slightly better than in IDH2 mutated AML without i(7)(p10), although this difference was not statistically significant. (5) Taking into account that the IDH2 mutation was always detected in the main clone and that no other targetable genes/aberrations were observed, these patients are potentially suitable for targeted therapy with IDH inhibitors.

Large Chromosome 2p Duplication-Associated Mechanisms and Clinical Presentations

Chromosome 2p (chr2p) duplication, also known as trisomy 2p, is a rare chromosome abnormality associated with developmental delay, intellectual disability, behavioral problems, and distinctive facial features. Most of the reported cases involving trisomy 2p include additional copy number variants (CNVs) in other regions of the genome and are usually small in size. Little is known about the clinical outcomes of large duplications of chr2p as the sole cytogenetic abnormality. In this study, 193 samples at the Greenwood Genetic Center (GGC) with CNVs involving chr2p were evaluated, out of which 86 had chr2p duplications. Among them, 8 patients were identified with large chr2p duplications ranging in size from 9.3 Mb to 89 Mb, and no deletions or duplications involving other chromosomes were identified in those patients. These duplications were associated with inverted duplication, tandem duplication, and duplication as the result of translocation, with no additional CNVs identified by microarray analysis. Confirmation by conventional cytogenetics was performed in 7 of the 8 patients, and the translocations were confirmed by fluorescence in situ hybridization. Interestingly, 1 patient was found to have mosaic complete trisomy 2p as the result of an unbalanced de novo (X;2) chromosomal translocation. X-inactivation was skewed toward the derivative X chromosome, yet it did not appear to extend into the chromosome 2 material. Various shared clinical manifestations were observed in the individuals in this study, including developmental delay, hemifacial hypoplasia, cleft palate, and short stature, and they also have distinct features such as hypotonia, cerebellar hypogenesis, and corpus callosum agenesis, which might result from a gene dosage effect of the duplication. In conclusion, single-event large chr2p duplications can result from different mechanisms, including inverted or tandem duplications within chromosome 2, or translocations involving chromosome 2 and other chromosomes. Partial or complete trisomy 2p is commonly associated with developmental delay, and additional clinical features may be related to gene dosage effects.

Therapeutic Outcomes of Relapsed-Refractory Multiple Myeloma Patients with 1q21+Treated with Daratumumab-Based Regimens: A Retrospective Analysis

Introduction: Gain/amplification of 1q21 (1q21+) is detected in approximately 40% of MM patients (pts) at diagnosis. There is emerging data that 1q21+ has a negative impact on prognosis in both newly diagnosed MM (NDMM) and relapsed/refractory MM (RRMM) pts even when treated with highly efficacious novel agents such as bortezomib (Bor), lenalidomide (Len), pomalidomide (Pom), carfilzomib (Car), and daratumumab (Dara). However, a recent pooled subgroup analysis of the phase 3 ICARIA-MM and IKEMA trials revealed that the addition of the isatuximab (Isa) to the backbone of Pom-dexamethasone (Dex) or Car-Dex, respectively, improved progression free survival (PFS) in RRMM pts with 1q21+. The benefit of Dara-based regimens in pts with RRMM harboring 1q21+ abnormalities is currently unknown. We evaluated the efficacy of Dara-based regimens in pts with RRMM harboring 1q21+. Methods: We identified consecutive MM pts who received a Dara-based regimen, between 1/2015 and 4/2019 at the Mayo Clinic Comprehensive Cancer Center's three sites (Rochester MN, Scottsdale AZ, Jacksonville FL). We included pts who had received any Dara-based regimen after 1-3 prior lines of therapy. 1q21+ abnormality was defined as gain of 1q21 [gain(1q21), 3 copies] and amplification (amp) of 1q21 [amp(1q21), ≥4 copies] on FISH. High-risk cytogenetics (HRCyto) was defined as FISH testing that was positive for t(4;14), t(14;16), t(14;20) or del(17p). Pt categories for analysis were: (1) with or without 1q21+, (2) standard-risk cytogenetics (SRCyto) vs. 1q21+ as the only cytogenetic abnormality, (3) HRCyto with or without 1q21+. Fisher's exact test was used to compare differences between categorial values and PFS was analyzed using Kaplan-Meier analysis. Results: 368 pts were identified, 278 had received a Dara-based regimen after 1-3 lines and 232 had FISH testing done and were included in the analysis. All pts received either Dara single agent, Dara-Len-Dex, Dara-Pom-Dex, or Dara-Bor-Dex. No pts received Dara-Car-Dex. Baseline characteristics are shown in Table 1. 1q21+ was noted in 57 pts (1q21 amp: 4, 1q21 gain: 53). The median follow-up time for the entire cohort was 35.7 months (m) (95% CI: 25.2-NR). The median PFS for 1q21+ pts treated with a Dara-based regimen was 24.6 m (95% CI: 19.9-29.4). The overall response rate (ORR) for pts with 1q+ treated with a Dara-based regimen was 57.9%; sCR: 1 (1.75%), CR: 3 (5.26%), VGPR: 9 (15.8%), and PR: 21 (36.8%). SD was noted in 16 (28%) pts. There were no PFS differences (p=0.956) or ORR differences (p=0.776) between the different Dara-based regimens used in pts with 1q21+. There were no PFS differences for pts with 1q21+ vs. pts without 1q21+; 24.5 m (95% CI: 18.6-29.4) vs. 23.5m (95% CI: 21.1-28.1), respectively; p=0.392. There were also no differences in ORR (p=0.09) for pts with 1q21+ vs. pts without 1q21+ treated with a Dara-based regimen. There were no PFS differences amongst pts with HRCyto without 1q21+ vs. pts with HRCyto and 1q21+; 23.5 m (95% CI: 20.7-NR) vs. 24.6 m (95% CI: 18.0-HR), respectively, p=0.507. Similarly, there were no ORR differences (p=0.127) between pts with HRCyto without 1q21+ vs. pts with HRCyto with 1q21+. Lastly, looking at pts with SRCyto vs. 1q21+ as the sole cytogenetic abnormality, there were no significant PFS differences [23.1 m (95% CI: 19.4-28.1), vs. 24.6 m (95% CI: 19.8-34.8), respectively, p=0.64] or ORR differences (p=0.075). Conclusion: Since 1q21+ is seen fairly frequently in pts with MM, its role as an independent prognostic marker as well as biomarker to guide treatment selection is being defined. While there is emerging data that Isa-based regimens can improve the PFS and ORR of patients with RRMM with 1q21+, we noted that for RRMM pts treated with Dara-based regimens, outcomes for pts with 1q21+ are similar to those with standard risk disease or other HRCyto markers. The lack of PFS and ORR differences seen regardless of 1q21+ status with Dara-based regimens could suggest that Dara-based treatment may abrogate the poor outcomes associated with the presence of 1q21+. To further explore the importance of 1q21+ as a therapeutic and prognostic biomarker in MM, existing data from large, randomized, prospective phase 3 trials needs to be pooled and analyzed to evaluate the role of anti-CD38 monoclonal antibodies in pts with 1q21+. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Rapid and Reproducible Karyotyping with Nanopore Sequencing in AML Patients

Introduction: We evaluated low coverage whole genome sequencing (WGS) of acute myeloid leukemia (AML) patients using long read Oxford Nanopore Technology (ONTseq) for karyotyping and compared the results and the frequency of patients with myelodysplasia-related cytogenetic abnormalities according to previous and current AML classifications with conventional cytogenetics (CG). Methods: Diagnostic samples from peripheral blood or bone marrow of 100 AML patients were sequenced in 3 medical centers on a GridION sequencer using SQK-LSK109 kits according to the manufacturer's instructions, and analyzed by CG. Ten samples were sequenced independently in two different laboratories. Fifty samples were sequenced prospectively from newly diagnosed de novo AML patients. Sequencing reads were analyzed by an in-house bioinformatics pipeline based on publicly available tools such as nglmr aligner, samtools, igv_tools, R package ACE, the CyDAS webserver, and custom scripts for transforming ACE copy number models to karyotype formulas. CNVs from ONTseq data were mapped to 290 chromosome bands to allow comparison with CG. We compared the following cytogenetic categories: Complex karyotype (cKT) ≥ 3 chromosomal abnormalities; monosomal karyotype (mKT) as defined by Breems et al.; karyotype with myelodysplasia-related cytogenetic abnormalities that defines MRC-AML according WHO 2016 classification (MRC-KT), and ICC 2022 classification (MRCA-KT). Results: ONTseq of 100 AML patients with a 4.4-fold genome coverage (range: 2.1-6.2) allowed us to resolve CNVs at a high resolution of ≥0.1 mega-basepairs (Mbp). Analysis of the ten duplicate samples established a high reproducibility of ONTseq. 110 CNVs were detected by both laboratories at identical chromosome positions and similar length (R2=0.999). Only 8 CNVs (7%) differed considerably in length between the replicates (median length difference 2 Mbp, range 0.4-16 Mbp). Of 100 patients with a median age of 65 years (range 18-97), ≥1 chromosome band was gained or lost in 69 patients according to ONTseq analysis, while CNVs or balanced translocations were found by cytogenetics in 72 patients. Five patients had a normal karyotype by ONTseq, but not by CG, while two patients had a normal karyotype by CG but not by ONTseq. Two of these 7 discrepant patients had translocations as their sole cytogenetic abnormality [t(8;21) and t(9;11)], which cannot be detected by low coverage genome ONTseq. Patients with chromosomal abnormalities had a median of 29 (range 1-140) and 30 (range 3-136) chromosomal bands with CNVs according to ONTseq and CG, respectively. Considering all patients, gains and losses at the level of chromosomal bands correlated very well (gains: R2=0.922; losses: R2=0.923) (Figure 1). All patients were then classified according the definitions for cKT, mKT, MRC-KT, and MRCA-KT (Table 1). Overall, patients were similarly classified by ONTseq and CG results (P<0.001). MRC and MRCA-KTs were consistently identified by ONTseq and CG in 57 patients. In 2 and 2 patients, MRC and MRCA-KTs, respectively, were identified by CG only. In 1 and 1 patient, MRC and MRCA-KT, respectively, was identified by ONTseq only. The largest discrepancies were found for mKTs due to the higher resolution of ONTseq, which more often identified subchromosomal instead of whole chromosome CNVs compared to CG, which were not considered for mKT. Fifty of the 100 patients were sequenced prospectively at time of diagnosis. The median age of these patients was 70 years (range 61-97) and all patients had de novo AML. At least one CNV was detected by ONTseq in 24 (48%) and by CG in 26 (52%) patients. cKT, mKT, MRC-KT, and MRCA-KT were detected by ONTseq in 12, 7, 17, and 17 patients and by CG in 14, 11, 17, and 17 patients, respectively. The median time for library preparation for prospectively analyzed patients was 3:45 hours, for ONTseq 24 hours and for bioinformatic analysis 6:30 hours. In this setting, clinical reports can be delivered on the fourth day of sample receipt. Conclusion: Low coverage WGS with ONT provides a highly reproducible method for rapid karyotyping with high resolution. The identification of cKT and myelodysplasia-related cytogenetic abnormalities correlated very well with conventional cytogenetics, even though balanced translocations cannot be detected by our ONTseq protocol. Low coverage WGS is a promising approach for diagnostic karyotyping in AML patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Genomic and clinical findings in myeloid neoplasms with PDGFRB rearrangement

Platelet-derived growth factor receptor B (PDGFRB) gene rearrangements define a unique subgroup of myeloid and lymphoid neoplasms frequently associated with eosinophilia and characterized by high sensitivity to tyrosine kinase inhibition. To date, various PDGFRB/5q32 rearrangements, involving at least 40 fusion partners, have been reported. However, information on genomic and clinical features accompanying rearrangements of PDGFRB is still scarce. Here, we characterized a series of 14 cases with a myeloid neoplasm using cytogenetic, single nucleotide polymorphism array, and next-generation sequencing. We identified nine PDGFRB translocation partners, including the KAZN gene at 1p36.21 as a novel partner in a previously undescribed t(1;5)(p36;q33) chromosome change. In all cases, the PDGFRB recombination was the sole cytogenetic abnormality underlying the phenotype. Acquired somatic variants were mainly found in clinically aggressive diseases and involved epigenetic genes (TET2, DNMT3A, ASXL1), transcription factors (RUNX1 and CEBPA), and signaling modulators (HRAS). By using both cytogenetic and nested PCR monitoring to evaluate response to imatinib, we found that, in non-AML cases, a low dosage (100–200 mg) is sufficient to induce and maintain longstanding hematological, cytogenetic, and molecular remissions.

Distinctive Flow Cytometric and Mutational Profile of Acute Myeloid Leukemia With t(8;16)(p11;p13) Translocation.

Acute myeloid leukemia (AML) with t(8;16)(p11;p13) abnormalities is a rare, aggressive, and diagnostically challenging subtype that results in KAT6A-CREBBP gene fusion. To investigate their immunophenotype and genomic features, we identified 5 cases of AML with t(8;16) through a retrospective review of the databases at Northwestern Memorial Hospital in Chicago, IL, and Washington University Medical Center, in St Louis, MO. In all, 4 of 5 cases were therapy related and 1 was possibly therapy related. The leukemic blasts showed distinctive features, including bright CD45 expression and remarkably high side scatter that overlapped with maturing myeloid elements, making the blasts difficult to identify on initial examination. They were positive for CD13, CD33, and CD64 and negative for CD34 and CD117. Next-generation sequencing profiling of 4 cases revealed pathogenic ASXL1 (2 cases), FLT3-tyrosine kinase domain (TKD) mutations (2 cases), and other pathogenic mutations. In 3 patients, t(8;16) was the sole cytogenetic abnormality; additional aberrations were found in 2 patients. Single nucleotide polymorphism microarray revealed 1 case with 7q deletion as a secondary clone. Our data highlight the distinctive immunophenotypic profile of AML with t(8;16), which, along with its unique morphology, often presents a diagnostic challenge. We showed that mutations of either ASXL1 or FLT3-TKD are seen in most cases of this leukemia.

Myeloid malignancies with translocation t(4;12)(q11-13;p13): molecular landscape, clonal hierarchy and clinical outcomes.

Translocation t(4;12)(q11‐13;p13) is a recurrent but very rare chromosomal aberration in acute myeloid leukaemia (AML) resulting in the non‐constant expression of a CHIC2/ETV6 fusion transcript. We report clinico‐biological features, molecular characteristics and outcomes of 21 cases of t(4;12) including 19 AML and two myelodysplastic syndromes (MDS). Median age at the time of t(4;12) was 78 years (range, 56–88). Multilineage dysplasia was described in 10 of 19 (53%) AML cases and CD7 and/or CD56 expression in 90%. FISH analyses identified ETV6 and CHIC2 region rearrangements in respectively 18 of 18 and 15 of 17 studied cases. The t(4;12) was the sole cytogenetic abnormality in 48% of cases. The most frequent associated mutated genes were ASXL1 (n = 8/16, 50%), IDH1/2 (n = 7/16, 44%), SRSF2 (n = 5/16, 31%) and RUNX1 (n = 4/16, 25%). Interestingly, concurrent FISH and molecular analyses showed that t(4;12) can be, but not always, a founding oncogenic event. Median OS was 7.8 months for the entire cohort. In the 16 of 21 patients (76%) who received antitumoral treatment, overall response and first complete remission rates were 37% and 31%, respectively. Median progression‐free survival in responders was 13.7 months. Finally, t(4;12) cases harboured many characteristics of AML with myelodysplasia‐related changes (multilineage dysplasia, MDS‐related cytogenetic abnormalities, frequent ASXL1 mutations) and a poor prognosis.

Clonal Evolution in CLL

Clonal Evolution in CLL

Impact of 9q Deletions on the Classification in AML

Background: In the 2016 revision of the World Health Organization (WHO) classification of myeloid neoplasms and acute leukemia the deletion of the long arm of chromosome 9 (del(9q)) was excluded as a defining cytogenetic abnormality for AML with myelodysplasia-related changes due to its frequent association with NPM1 and biallelic CEBPA mutations.Aim: Investigation of the frequency of del(9q) in AML and its accompanying molecular and cytogenetic abnormalities to assess whether or not del(9q) is associated with a myelodysplasia-related mutation profile.Patients and Methods: We evaluated 9762 AML patients (pts) diagnosed between 2005 and 2017 for del(9q) using chromosome banding analysis (CBA). Pts with del(9q) were further analyzed for mutations in NPM1 , CEBPA and RUNX1 with amplicon sequencing to categorize them according to the WHO classification. Pts without a class defining aberration or a complex aberrant karyotype were screened for AML- or MDS-related mutations (ASXL1, BCOR, EZH2, FLT3- ITD , FLT3- TKD , SF3B1, SRSF2, STAG2, TET2, TP53, U2AF1, WT1 and ZRSR2 ). Variants of unknown significance were excluded from statistical analysis. The size of the deleted region on chromosome 9 was evaluated by array CGH (Agilent Technologies, Santa Clara, CA) and NUP98 -rearrangements were investigated by FISH(MetaSystems, Altlussheim, Germany).Results: In 114/9762 pts (1 %) a del(9q) was detected by CBA. This cohort with del(9q) comprised 60 male and 54 female pts, median age was 63 years (range: 16 - 90 years). Del(9q) was accompanied by the following rearrangements: RUNX1-RUNX1T1 rearrangement in 20/114 pts (18 %), PML-RARA in 5/114 pts (4 %), KMT2A in 2/114 pts (2 %) and MECOM in 1/114 pts (1 %). In 20/114 pts (18 %) a NPM1 mutation and in 7/114 pts (6 %) a biallelic CEBPA mutation was observed. Furthermore, RUNX1 mutations were found in 3/114 pts (3 %). The remaining 56 pts (49 %) were divided into the following three groups: pts with del(9q) as a sole cytogenetic abnormality (del(9q) sole, 8/114 pts, 7 %), pts with one or two additional cytogenetic aberrations (del(9q) other, 19/114 pts, 17 %) and pts with complex aberrant karyotype (complex, 29/114 pts, 20 %) harboring 4 to 29 cytogenetic aberrations (median: 8). Cases with del(9q) sole and other (n = 27) were further characterized. NUP98 -FISH analysis revealed a NUP98 -rearrangement in 5/27 pts (19 %) in the del(9q) sole and other group. Moreover, two pts with del(9q) showed AML specific rearrangements (t(8;16)(p11;p13) and t(3;5)(q25;q35)). The most frequently mutated genes in del(9q) sole and other were: FLT3 -ITD (8/23 pts, 35 %), TET2 (7/27 pts, 26 %), FLT3 -TKD (4/21 pts, 19 %) and WT1 (3/26 pts, 11 %). A very high frequency of FLT3 -ITD and FLT3 -TKD were found in pts with del(9q) and balanced rearrangements (5/7 pts, 71 %).To identify whether pts with del(9q) sole and other rather resemble a molecular mutation profile typical for secondary AML arising from MDS (s-AML), the presence of gene mutations related to s-AML (according to Lindsley, et al., 2015, Blood) was evaluated. 9 pts (33 %) had a mutation in at least one of these genes, 14 pts (52 %) showed no such mutation and 4 pts (15 %) could not be fully analyzed.Array CGH showed that in pts with del(9q) sole the deletion size ranged from 14 - 42 Mb (median: 28 Mb), in pts with del(9q) other from 19 - 70 Mb (median: 32 Mb). A minimal deleted region for pts with del(9q) sole and other could not be determined for all pts due to the strong variation between the respective deleted regions.Follow up data was available for 75 pts with a median follow-up of 27 months. The median overall survival (OS) for pts with RUNX1-RUNX1T1 , NPM1 and bialleic CEBPA mutation was not reached. It was 73 month for pts with del(9q) sole and other and 16 month for pts with complex karyotype, RUNX1 mutations or MECOM rearrangements. The OS of first two groups did not differ significantly, but was significantly distinct from the last group (p = 0.03 and 0.03, respectively).Conclusion: In 1 % of AML pts a del(9q) was present. Del(9) frequently co-occured with RUNX1-RUNX1T1 , biallelic CEBPA and NPM1 mutations, NUP98 -rearrangements and other AML-typical translocations. A mutation signature typical for s-AML was infrequent. Thus, it seems reasonable that the del(9q) is no longer regarded as a defining cytogenetic abnormality for AML with myelodysplasia-related changes, in particular as prognosis in del(9q) cases with non-complex karyotype is favorable. DisclosuresBalk:MLL Munich Leukemia Laboratory: Employment. Stengel:MLL Munich Leukemia Laboratory: Employment. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Fasan:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

38. A rare constitutional 13q deletion associated with retinoblastoma and chronic lymphocytic leukemia

The 13q deletion syndrome is a rare congenital disorder characterized by a wide phenotypic spectrum depending on the size and location of the deletion, with clinical features spanning intellectual disability, craniofacial dysmorphism, congenital malformations, and retinoblastoma (Mitter et al., 2011). In the somatic setting, deletion of 13q14 is the most common cytogenetic abnormality in chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), and is associated with a favorable outcome when present as the sole cytogenetic abnormality (Van Dyke et al., 2010). To date, published reports and review of the literature have not associated constitutional 13q deletions to an increased risk of CLL/SLL. Here, we describe the molecular and cytogenetic analysis of a 47-year-old male referred to our laboratory for CLL genetic analysis. By fluorescence in situ hybridization (FISH), peripheral blood nuclei exhibited a homozygous 13q14 deletion in ∼63% of nuclei, and a heterozygous deletion in the remaining ∼37% of nuclei. FISH also identified heterozygous loss of the RB1 gene region in ∼97% of nuclei, consistent with the patient's history of retinoblastoma. Conventional chromosome analysis identified a large del(13)(q14.1q21.2), and chromosome microarray (CMA) mapped the deletion to 13:43,773,101-69,897,491 (26.1Mb, hg19) in a perfect heterozygous state. In essential agreement with the FISH results, CMA also identified a homozygous 796Kb deletion including MIR16-1, MIR15A, and DLEU1/2 in ∼50% of cells; this homozygous deletion likely represents the CLL cell population. To our knowledge this is the first case of a constitutional 13q deletion associated with adult CLL. Given that monoallelic deletions of RB1 constitute ∼6% of RB1’s mutational spectrum (Albretch et al., 2005; Castéra et al., 2013), a better understanding of this patient may help explore the risk for adult-onset CLL in those individuals with an infantile history of retinoblastoma who carry an RB1 deletion rather than a point mutation.

Favourable outcomes for high-risk Burkitt lymphoma patients (IPI 3-5) treated with rituximab plus CODOX-M/IVAC: Results of a phase 2 UK NCRI trial.

IntroductionOutcomes after frontline treatment of Burkitt lymphoma (BL) have improved with the introduction of dose‐intense chemotherapy regimens, such as CODOX‐M/IVAC. While rituximab has increased survival rates for most forms of high‐grade B‐cell lymphoma, there has previously been hesitancy about incorporating it into BL treatment, partly due to concerns about increased toxicity. Prospective data using the standard dose CODOX‐M/IVAC regimen in combination with rituximab are lacking. We conducted a single‐arm phase 2 trial to assess the efficacy and toxicity of R‐CODOX‐M/R‐IVAC.MethodsEligible patients were aged 18–65 years, with newly diagnosed BL with MYC rearrangement as the sole cytogenetic abnormality, and high‐risk disease, defined by an International Prognostic Index (IPI) score of 3‐5. Patients received two cycles of R‐CODOX‐M chemotherapy alternating with two cycles of R‐IVAC, followed by two further cycles of rituximab alone. The primary endpoint was 2‐year progression‐free survival.ResultsThirty‐eight patients were registered but after central pathology review, 27 patients had confirmed BL and commenced study treatment. Median age was 35 years, 14.8% patients had central nervous system involvement and 18.5% were HIV positive. Twenty‐two (81.4%) patients completed four cycles of chemotherapy. There were two treatment‐related deaths (7.4%). Two‐year progression‐free and overall survival rates were 77.2% (90% confidence interval [CI]: 56.0‐89.0) and 80.7% (90% CI: 59.6‐91.5), respectively.ConclusionsThis prospective trial demonstrates excellent survival rates with R‐CODOX‐M/R‐IVAC in a high‐risk BL cohort. It provides reassuring evidence regarding the feasibility of this regimen and also provides a benchmark for future studies.

Features of an Unusual AML with t(4;12)/Pdgra-ETV6 As the Sole Cytogenetic Abnormality Associated with Myelodysplastic Changes: A Case Report and Literature Review

The t(4;12) cytogenetic abnormality is rare in AML, and is associated with a unique morphologic and immunophenotypic profile. Pseudo-lymphoid morphology of myeloid blasts (appearing as atypical lymphocytes), multilineage dysplasia, absence of eosinophilia, aberrant CD7 and lack of myeloperoxidase (MPO) expression, low frequency of FLT3 mutations, and treatment refractoriness are some of the characteristics of this AML subtype previously described in the literature. We report a case of AML with t(4;12) in a middle-age male who presented with pancytopenia: WBC 3.34 (65% lymphocytes), Hgb 6.9, Plt 13, and no known prior history of a myeloid neoplasm. Bone marrow aspirate revealed 78% blasts that appeared large with uneven chromatin distribution, prominent nucleoli, and inconspicuous granules; and significant dysplastic features in the erythroid and megakaryocyte elements. Immunostains performed on the trephine core biopsy showed blasts expressed CD34 (subset), CD71 (subset), and were CD117+ Bcl-2+ CD43+ CD7+ MPO-. Flow cytometry identified 74% myeloblasts, positive for CD34, CD38, CD123, CD117, CD33, HLA-DR, CD13 (subset), CD56, CD7, CD71; negative for monocyte-specific markers. The morphologic and immunophenotypic features were compatible with AML with MDS-related changes. Karyotype analysis identified t(4;12)(q12;p13) in 18/20 cells, and no additional chromosomal abnormalities. PDGFRA translocation to 12p was confirmed using PDGFRA and ETV6 FISH probes. Analysis of frequent genomic alterations in AML identified BCOR p.Leu1646Profs, TET2 p.Gln1627Ilefs, U2AF1 p.Ser34Phe mutations; and no FLT3 mutation. Conclusions: In AML with t(4;12)/PDGFRA-ETV6, though lack of a FLT3 mutation is relatively favorable, BCOR and U2AF1 mutations and multilineage dysplasia are unfavorable prognostic factors. Synergism of the additional oncogenic mutations with the PDGFRA activating mutation can be postulated. The t(4;12)/PDGFRA-ETV6 should be recognized in the WHO classification as a recurrent balanced translocation in AML with MDS-related changes. Disclosures No relevant conflicts of interest to declare.

Outcomes of Patients with Limited-Stage Aggressive Large B-Cell Lymphoma with MYC Rearrangement with and without BCL2 and/or BCL6 Rearrangements: A Retrospective Analysis from 15 US Academic Centers

Outcomes of Patients with Limited-Stage Aggressive Large B-Cell Lymphoma with MYC Rearrangement with and without BCL2 and/or BCL6 Rearrangements: A Retrospective Analysis from 15 US Academic Centers

3q26/EVI1 rearrangement in myelodysplastic/myeloproliferative neoplasms: An early event associated with a poor prognosis

3q26.2/EVI1 rearrangements resulting in EVI1 overexpression play an important role in leukemogenesis and are associated with treatment resistance and a poorer prognosis in patients with acute myeloid leukemia, myelodysplastic syndrome, chronic myeloid leukemia and BCR-ABL negative myeloproliferative neoplasms. In this study, we aim to explore the clinicopathological features of myelodysplastic/myeloproliferative (MDS/MPN) neoplasms with 3q26.2/EVI1 rearrangements and determine the potential impact of these cytogenetic abnormalities on treatment response and survival. The study group included 12 cases of MDS/MPN with 3q26.2 rearrangements detected by conventional karyotyping. There were 7 men and 5 women with a median age of 67 years (range, 51–79 years) at time of initial MDS/MPN diagnosis. Ten cases were classified as chronic myelomonocytic leukemia (CMML) and 2 were MDS/MPN, unclassifiable. Among CMML cases, 5 (50%) were proliferative type and 5 (50%) were dysplastic type. Based on blast counts, these 10 CMML were: CMML-0 (n = 2), CMML-1 (n = 3), and CMML-2 (n = 5). Eleven (92%) patients had 3q26 rearrangements at the initial diagnosis. Inv(3)(q21q26.2) was most common, identified in 7(58%) patients, followed by t(3;21)(q26.2;q22) in 2 patients and 1 patient each with t(3;3)(q21;q26.2), t(2;3)(p21;q26-27), and t(3;6)(q26.2;q26). Six (50%) patients had 3q26.2 rearrangements as a sole cytogenetic abnormality and 6 (50%) patients had additional cytogenetic abnormalities. Molecular studies revealed DNMT3A mutations in all 3 patients assessed and RAS mutations in 2 of 8 (25%) patients. No mutations in ASXL1 (n = 3), TET2 (n = 3), FLT3 ITD/D835 (n = 10), and CEBPA (n = 7) were detected. Most patients received hypomethylating agent based chemotherapy. The median follow-up was 11.5 months (range, 1.5–24 months) and at time of last follow-up, 11 (92%) died with a median survival of 13.4 months (range, 1.5–24 months). The only patient alive had a relatively short follow-up of 2.4 months and showed disease progression at the last visit. In conclusion, 3q26.2/EVI1 rearrangements are a rare event and usually present at time of initial diagnosis in MDS/MPN. The presence of 3q26.2/EVI1 rearrangements in MDS/MPN is associated with rapid disease progression, poor response to treatment, and a poor prognosis.

Combining cytogenetic and epigenetic approaches in chronic lymphocytic leukemia improves prognosis prediction for patients with isolated 13q deletion

BackgroundBoth defective DNA methylation and active DNA demethylation processes are emerging as important risk factors in chronic lymphocytic leukemia (CLL). However, associations between 5-cytosine epigenetic markers and the most frequent chromosomal abnormalities detected in CLL remain to be established.MethodsCLL patients were retrospectively classified into a cytogenetic low-risk group (isolated 13q deletion), an intermediate-risk group (normal karyotype or trisomy 12), and a high-risk group (11q deletion, 17p deletion, or complex karyotype [≥ 3 breakpoints]). The two 5-cytosine derivatives, 5-methylcytosine (5-mCyt) and 5-hydroxymethylcytosine (5-hmCyt), were tested by ELISA (n = 60), while real-time quantitative PCR was used for determining transcriptional expression levels of DNMT and TET (n = 24).ResultsBy using global DNA methylation/demethylation levels, in the low-risk disease group, two subgroups with significantly different clinical outcomes have been identified (median treatment-free survival [TFS] 45 versus > 120 months for 5-mCyt, p = 0.0008, and 63 versus > 120 months for 5-hmCyt, p = 0.04). A defective 5-mCyt status was further associated with a higher percentage of 13q deleted nuclei (> 80%), thus suggesting an acquired process. When considering the cytogenetic intermediate/high-risk disease groups, an association of 5-mCyt status with lymphocytosis (p = 0.0008) and the lymphocyte doubling time (p = 0.04) but not with TFS was observed, as well as a reduction of DNMT3A, TET1, and TET2 transcripts.ConclusionsCombining cytogenetic studies with 5-mCyt assessment adds accuracy to CLL patients’ prognoses and particularly for those with 13q deletion as a sole cytogenetic abnormality.

The prognostic significance of 13q deletions of different sizes in patients with B-cell chronic lymphoproliferative disorders: a retrospective study.

Patients with chronic lymphocytic leukemia (CLL) with 13q deletion as the sole cytogenetic abnormality usually have a favorable outcome, but the frequency of the 13q14 deletion and its impact on the outcome of other B-cell chronic lymphoproliferative disorders (BCLPDs) remain unclear. To further characterize this aberration, we investigated the prognostic significance of 13q deletion in 541 patients with BCLPDs. We performed fluorescence in situ hybridization (FISH) studies with 13q locus-specific LSI-D13S25 and LSI-RB1 probes. 52.1% of the patients with CLL harbored 13q deletion, which was significantly higher than that of other BCLPDs (p<0.001). The size of 13q deletion was heterogeneous in both CLL and other BCLPDs. However, the distribution of cases with different deletion sizes showed no significant difference between the two groups. Whereas 13q deletion was a favorable prognostic factor in CLL, a large deletion of 13q was associated with poor prognosis in terms of time to first therapy (p=0.020), progression-free survival (p=0.05) and overall survival (p=0.002) in BCLPD cases other than CLL. In conclusion, the deletion of 13q varied in size both in CLL and in other BCLPDs and adversely influenced the prognosis of patients with other BCLPDs.

Analyzing the Transcriptome Discovers up-Regulation of HOXA Genes in Patients with Myeloid Neoplasms and Isochromosome 17q and Mutations in ASXL1, SETBP1 and SRSF2

Introduction: Isochromosome 17 (i(17q)) is a rare cytogenetic abnormality reported in different myeloid neoplasms. I(17q) has been described as primary and as secondary chromosomal aberration, often acquired in the disease course. Recently, we have shown that patients with i(17q) show a distinct mutation profile with mutations in ASXL1, SETBP1 and SRSF2. Further data suggested a parallel acquisition of SETBP1 mutation and i(17q) (Meggendorfer et al., Leukemia, 2016). Of note, i(17q) results in three copies of the splicing factor SRSF2, potentially further influencing the transcriptome of affected cells.Aim: To characterize the transcriptome in myeloid neoplasms with i(17q) and the typical mutations in ASXL1, SETBP1 and SRSF2 (A/S/S/i17pos).Patients and Methods: In total 18 patients were selected based on the cytogenetic profile and the molecular mutations. All had the diagnosis of a myeloid neoplasm by cytomorphology according to the WHO. Chromosome banding and FISH analysis and mutation status of ASXL1, SETBP1 and SRSF2 was available in all cases. Three patient groups were defined: 1) ASXL1, SETBP1, SRSF2 mutated and a normal karyotype (A/S/S/-pos; n=5) and gain of i(17q) during follow up, 2) ASXL1, SETBP1, SRSF2 mutated and i(17q) as sole cytogenetic abnormality (A/S/S/i17pos; n=8), 3) ASXL1, SRSF2 mutated and i(17q) as sole cytogenetic abnormality (A/-/S/i17pos; n=5). In all cases RNA sequencing was performed (TruSeq RNA Sample Preparation V2, Illumina, San Diego, CA). A control RNA (Universal Human Reference RNA, Agilent Technologies, Santa Clara, CA) was investigated in triplicate for normalization and comparison. Expression analyses were performed with BaseSpace RNA Express app (Illumina, San Diego, CA).Results: In total 23,710 genes were annotated by RNA sequencing and at least 14,773 analyzed for differential expression, showing that all three patient groups show aberrant expression in comparison to the control RNA. In mean 4,930 genes/group (range: 3,603-6,711) were differentially expressed. The differential expression ranged from -12.7 to 11.2 log2(fold change). Comparing the two groups with all three gene mutations but different i(17q) status (A/S/S/-pos and A/S/S/i17pos) showed that the presence of i(17q) changes the expression pattern with 1,596/13,218 assessed genes differentially expressed. In detail, in A/S/S/i17pos cases 790 genes were significantly over expressed while 806 genes showed reduced expression compared to A/S/S/-pos, ranging from -2.84 to 2.78 log2(fold change). The expression of SRSF2 was not affected by i(17q), although i(17q) cases show three SRSF2 gene copies. Analyzing the most strongly affected genes (2<log2(fold change)<-2, n=48) showed that differential gene expression affected mostly the homeobox (HOX) genes clustering on chromosome 7p15 (HOXA1, HOXA5 and HOXA7) as well as MEIS1, an important cofactor of HOX genes. HOX genes represent a family of transcription factors, shown to be involved in hematopoiesis. Addressing specifically the differential expression of HOX genes showed that further 4 HOXA genes (HOXA2, HOXA4, HOXA9, HOXA10) and 3 HOXB genes (HOXB2, HOXB3, HOXB4) were significantly dysregulated, with HOXB clustering on 17q21. The expression of all HOX genes was up-regulated in cases with i(17q). Interestingly, the molecular mutation pattern A/S/Spos has also been shown to associate with patients having a monosomy 7 (Meggendorfer, #1364, ASH 2013), where the HOXA gene cluster is located. Comparing the HOXA and MEIS1 gene expression in all 18 samples to human control RNA revealed a significant lower expression level in A/S/S/-pos and an increased one in A/S/S/i17pos and A/-/S/i17pos patients, clearly differentiating i(17q) carrying patients. However, the additional SETBP1 mutation did not influence the expression pattern as seen by comparing A/S/S/i17pos and A/-/S/i17pos patients (0/13,398 assessed genes differentially expressed).Conclusion: 1) Transcriptome analysis of patients with myeloid malignancies, i(17q) and mutations in ASXL1, SETBP1 and SRSF2 show an up-regulation of HOXA genes. 2) Accompanying SETBP1 mutation does not further influence the transcriptome of A/-/S/i17pos patients. 3) The up-regulation of HOXA genes might indicate a pathogenic mechanism in patients with ASXL1, SETBP1, SRSF2 and i(17q). However, this finding has to be validated in a larger cohort and with an independent method. DisclosuresMeggendorfer:MLL Munich Leukemia Laboratory: Employment. Nadarajah: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.

Identification of a Novel Chromosomal Translocation t(11;16)(q23;q22) Fusing MLL to Enhancer of mRNA Decapping (EDC)-4 in Smoldering Acute Myeloid Leukemia

Abstract Background: Translocations involving the MLL gene located on chromosome 11q23 are usually found in de novo acute myeloid leukemia (AML) and generally confer a dismal prognosis unless the AF9 gene is involved (Döhner et al., Blood 2010;115:453-74). MLL can be fused to multiple different genes, resulting in the large and growing "MLL recombinome" (Meyer et al., Leukemia 2013;27:2165-76). Thus far, only two genes encoding proteins that are part of the mRNA decapping protein complex (i.e., DCP1A, DCPS) have been described as rarely being fused to MLL. Here, we describe an AML with an indolent disease course arising from myelodysplastic syndrome (MDS) that disclosed a unique MLL fusion with another component of the mRNA decapping complex, i.e., EDC4. Patient and Methods: Briefly, a 55 year-old female patient presented with an MDS [timepoint (t) -1] that within 10 months progressed to an AML with 2% blood and 40% bone marrow myeloblasts (t0). The patient refused treatment beyond supportive care. Six months later, a marked blast expansion of 80% was detectable in the blood (t1). The patient received 5 cycles of decitabine (t2, cycle 5), followed by 3 months of hydroxyurea (t3). Samples were depleted from CD3+ cells via MACS; CD3+ cells served as germline control. RNA sequencing libraries were prepared using the NEBNext Ultra RNA Library Prep Kit for Illumina (New England Biolabs), and the SureSelect Human All Exon v5 kit (Agilent Technologies) was used for exome capturing from gDNA. Next generation sequencing was performed on an Illumina Hiseq 2500. The analyses were performed as previously described (Kataoka et al. Nature Genetics 2015;47:1304-15, Becker et al. Blood 2014;123:1883-6). NOD scid gamma mice were used as hosts for patient derived tumor xenografts (PDX). Results: Standard metaphase cytogenetics at the diagnosis of AML (t0) revealed a previously undescribed translocation involving the MLL gene, i.e., t(11;16)(q23;q22), as the sole cytogenetic abnormality. The unknown fusion partner on chromosome 16 was identified by RNA-sequencing as the EDC4 gene (a.k.a. Ge-1), which encodes a key scaffold protein of the mRNA decapping complex; the fusion was confirmed by PCR on cDNA. The translocation led to the in-frame fusion of MLL exon 13 to EDC4 exon 6 which was linked by 19 nucleotides from EDC4 intron 5. The predicted amino acid sequence of the linker was ALNTLLR. Further analyses including exome sequencing on the samples collected over the disease course demonstrated STAG2 as a potential founder mutation that was already present during the MDS (t-1) and persisted throughout the disease course at variant allele frequencies (VAFs) of approximately 45-50%. At the time of transformation to AML (t0), the MLL-EDC4 and two RAS mutations (KRAS p.G13D, NRAS p.G12C) were detectable. Towards the terminal phase (t3), the RAS mutations disappeared and a clone that acquired a mutation in the FLT3 tyrosine kinase domain (TKD; p.D835V; VAF 43%) expanded. Primary blasts from the patient engrafted in NOD scid gamma mice and established a stable PDX serial transplantation mouse model used for drug testing. Conclusions: This report provides the first demonstration of an MLL-EDC4 in-frame fusion, with potential cooperativity with a founder mutation in the STAG2 splicing factor gene during the transformation of MDS to AML and the additional acquisition of a FLT3-TKD mutation during disease progression. RNA sequencing proved to be a very feasible approach to identify novel fusion partners of known oncogenes such as MLL. Disclosures Becker: BMS: Honoraria; Novartis: Honoraria. Kataoka:Yakult: Honoraria; Boehringer Ingelheim: Honoraria; Kyowa Hakko Kirin: Honoraria. Schüler:Oncotest GmbH: Employment. Ogawa:Kan research institute: Consultancy, Research Funding; Sumitomo Dainippon Pharma: Research Funding; Takeda Pharmaceuticals: Consultancy, Research Funding. Lübbert:Janssen-Cilag: Other: Travel Funding, Research Funding; Celgene: Other: Travel Funding; Ratiopharm: Other: Study drug valproic acid.