Abnormalities In Acute Myeloid Leukemia Research Articles

Immunotherapeutic Potential of Mutated NPM1 for the Treatment of Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is a malignant disease of the blood and bone marrow that is characterized by uncontrolled clonal proliferation of abnormal myeloid progenitor cells. Nucleophosmin 1 (NPM1) gene mutations are the most common genetic abnormality in AML, detectable in blast cells from about one-third of adults with AML. AML NPM1mut is recognized as a separate entity in the World Health Organization classification of AML. Clinical and survival data suggest that patients with this form of AML often have a more favorable prognosis, which may be due to the immunogenicity created by the mutations in the NPM1 protein. Consequently, AML with NPM1mut can be considered an immunogenic subtype of AML. However, the underlying mechanisms of this immunogenicity and associated favorable survival outcomes need to be further investigated. Immune checkpoint molecules, such as the programmed cell death-1 (PD-1) protein and its ligand, PD-L1, play important roles in leukemogenesis through their maintenance of an immunosuppressive tumor microenvironment. Preclinical trials have shown that the use of PD-1/PD-L1 checkpoint inhibitors in solid tumors and lymphoma work best in novel therapy combinations. Patients with AML NPM1mut may be better suited to immunogenic strategies that are based on the inhibition of the PD-1 immune checkpoint pathway than patients without this mutation, suggesting the genetic landscape of patients may also inform best practice for the use of PD-1 inhibitors.

Significance of targeting DNMT3A mutations in AML.

Acute myeloid leukemia (AML) is the most prevalent form of leukemia among adults, characterized by aggressive behavior and significant genetic diversity. Despite decades of reliance on conventional chemotherapy as the mainstay treatment, patients often struggle with achieving remission, experience rapid relapses, and have limited survival prospects. While intensified induction chemotherapy and allogeneic stem cell transplantation have enhanced patient outcomes, these benefits are largely confined to younger AML patients capable of tolerating intensive treatments. DNMT3A, a crucial enzyme responsible for establishing de novo DNA methylation, plays a pivotal role in maintaining the delicate balance between hematopoietic stem cell differentiation and self-renewal, thereby influencing gene expression programs through epigenetic regulation. DNMT3A mutations are the most frequently observed genetic abnormalities in AML, predominantly in older patients, occurring in approximately 20-30% of adult AML cases and over 30% of AML with a normal karyotype. Consequently, the molecular underpinnings and potential therapeutic targets of DNMT3A mutations in AML are currently being thoroughly investigated. This article provides a comprehensive summary and the latest insights into the structure and function of DNMT3A, examines the impact of DNMT3A mutations on the progression and prognosis of AML, and explores potential therapeutic approaches for AML patients harboring DNMT3A mutations.

WNT inhibitory factor 1 (WIF1) is a novel fusion partner of RUNX family transcription factor 1 (RUNX1) in acute myeloid leukemia with t(12;21)(q14;q22).

As a member of the core transcription factor family, RUNX1 plays an important role in stem cell differentiation. RUNX1 rearrangements are common in myeloid and lymphoid tumors [1]. (Blood 129(15):2070-2082, 2017). One of the most commonly detected abnormalities in acute myeloid leukemia (AML) is the translocation t(8;21)(q22;q22) (Blood Adv 4(1):229-238, 2020), resulting in a RUNX1::RUNX1T1 fusion. Occasionally, RUNX1 is translocated with other genes. This article describes an AML patient with a specific chromosomal translocation involving the RUNX1 gene and the identification of the RUNX1::WIF1 fusion. Chromosomal abnormalities were detected through karyotype analysis, break gene involved was identified via fluorescence in situ hybridization (FISH), and the novel fusion was identified through transcriptome sequencing and subsequently confirmed through reverse transcription-polymerase chain reaction (RT-PCR) and Sanger sequencing. A 79-year-old female patient diagnosed with AML was found to have a t(12;21)(q14;q12) translocation. FISH analysis provided evidence of RUNX1 gene rearrangement. Additionally, transcriptomic sequencing revealed a novel fusion known as RUNX1::WIF1, which consists of RUNX1 exon 2 and WIF1 exon 3. The novel fusion was further confirmed through RT-PCR and Sanger sequencing. We identified WIF1 as a novel fusion partner of RUNX1 in AML. Additionally, this is the first report of a RUNX1 fusion gene with the break point in intron 2, resulting in an out-of-frame fusion. Further research is needed to investigate the impact of this novel fusion on the establishment and progression of the disease.

Rearrangements involving 11q23.3/KMT2A in adult AML: mutational landscape and prognostic implications – a HARMONY study

Balanced rearrangements involving the KMT2A gene (KMT2Ar) are recurrent genetic abnormalities in acute myeloid leukemia (AML), but there is lack of consensus regarding the prognostic impact of different fusion partners. Moreover, prognostic implications of gene mutations co-occurring with KMT2Ar are not established. From the HARMONY AML database 205 KMT2Ar adult patients were selected, 185 of whom had mutational information by a panel-based next-generation sequencing analysis. Overall survival (OS) was similar across the different translocations, including t(9;11)(p21.3;q23.3)/KMT2A::MLLT3 (p = 0.756). However, independent prognostic factors for OS in intensively treated patients were age >60 years (HR 2.1, p = 0.001), secondary AML (HR 2.2, p = 0.043), DNMT3A-mut (HR 2.1, p = 0.047) and KRAS-mut (HR 2.0, p = 0.005). In the subset of patients with de novo AML < 60 years, KRAS and TP53 were the prognostically most relevant mutated genes, as patients with a mutation of any of those two genes had a lower complete remission rate (50% vs 86%, p < 0.001) and inferior OS (median 7 vs 30 months, p < 0.001). Allogeneic hematopoietic stem cell transplantation in first complete remission was able to improve OS (p = 0.003). Our study highlights the importance of the mutational patterns in adult KMT2Ar AML and provides new insights into more accurate prognostic stratification of these patients.

Small Molecule Menin Inhibitors: Novel Therapeutic Agents Targeting Acute Myeloid Leukemia with KMT2A Rearrangement or NPM1 Mutation.

Recent advances have included insights into the clinical value of genomic abnormalities in acute myeloid leukemia (AML) and consequently the development of numerous targeted therapeutic agents that have improved clinical outcome. In this setting, various clinical trials have recently explored novel therapeutic agents either used alone or in combination with intensive chemotherapy or low-intensity treatments. Among them, menin inhibitors could represent a novel group of targeted therapies in AML driven by rearrangement of the lysine methyltransferase2A (KMT2A) gene, previously known as mixed-lineage leukemia (MLL), or by mutation of the nucleophosmin1 (NPM1) gene. Recent phase1/2 clinical trials confirmed the efficacy of SNDX-5613 (revumenib) and KO-539 (ziftomenib) and their acceptable tolerability. Several small molecule menin inhibitors are currently being evaluated as a combination therapy with standard of care treatments. The current paper reviews the recent progress in exploring the inhibitors of menin-KMT2A interactions and their application prospects in the treatment of acute leukemias.

C-Kit and Flt3 mutation status in acute myeloid leukaemia with cytogenetic correlation and prognosis: A series of 75 cases

Recent WHO classification (5 edition) has included Acute Myeloid Leukaemia (AML) with mutations in NPM1, CEBPA as separate entities along with AML with cytogenetic abnormalities in AML with recurrent genetic abnormalities. Even though C-kit and FLT3 mutations in AML have no diagnostic importance, prognostic significance in different subtypes of AML for the presence of these mutations are not the same.To assess the correlation between French American British (FAB) AML classification, specific cytogenetic abnormalities with C-kit, and FLT3 mutation in AML and to evaluate the prognosis and survival of AML patients with respect to cytogenetic abnormalities and C-kit and FLT3 mutation status. Retrospectively all AML cases in which C-kit and FLT3 mutation status was assessed were retrieved; C-kit D816V mutation status had been assessed by Real time PCR; For FLT3 mutation, both Internal tandem duplication (ITD) and D835V mutation status had been assessed using PCR and gel electrophoresis; The data regarding morphological with immunophenotypic diagnosis, conventional karyotyping, FISH for translocation 8;21 (t (8;21)) and inversion16 / translocation16;16 (t (16;16) were also retrieved in all these cases along with follow-up from hospital records. Total 75 cases were included; Male/ Female ratio was 1.21:1 (41/34); Median age was 31 (Range: 2 - 64); 18 cases had translocation 8;21 (t (8;21)); 3 cases showed inversion16 / translocation16;16 (t (16;16); Out of the 18 cases which showed t (8;21), 10 cases had associated loss of sex chromosome. Eight cases had C-kit D816V mutation; three of which had t (8;21) while two had inversion 16. 12 cases had FLT3 mutations among which nine were ITD while three had D835V mutation. On karyotyping, one of these cases showed hyperdiploidy while the majority had normal karyotype. A single case had both C-kit D816V mutation and FLT3 D835V mutation with inversion 16 on karyotyping; Most common type of AML in both cases with FLT3 mutation and C-kit mutation was AML-M2 (FAB); Commonest karyotyping abnormality for cases with C-kit mutation was t(8;21); while for FLT3 mutation, the majority had normal karyotype; The single case which had both C-kit D816V mutation and FLT3 D835V mutation was alive event-free at three-year follow-up. Both FLT3 ITD and TKD mutations had a worse prognosis in our study. However, AML cases with C-kit mutation had a similar prognosis comparable to C kit negative cases. A large-scale study is required to elucidate the significance of this.

Encouraging Outcome of AML with t(8;16) Following Allogeneic Stem Cell Transplantation in First Remission - a Retrospective Analysis on Behalf of the Acute Leukemia Working Party of the EBMT

In the 2022 European Leukemia Net (ELN) risk classification, acute myeloid leukemia (AML) with translocation t(8;16) was newly classified as AML defined by specific cytogenetic abnormalities, associated with unfavorable prognosis. Translocation (8;16)(p11;p13) is a rare abnormality in AML with female preponderance and occurs more frequently in secondary AML (sAML), especially therapy-related, than in de novo disease. Clinically, AML with t(8;16) is associated with high risk of bleeding, occurrence of extramedullary manifestations, myelomonocytic differentiation, and signs of hemophagocytosis in the bone marrow. The translocation leads to the fusion of acetyltransferase KAT6A (also known as MOZ or MYST3) and CREB binding protein ( CREBBP) genes, both of them involved in hematopoiesis. ELN risk stratification of AML with t(8;16) was based on studies that included relatively low numbers of patients receiving allogeneic stem cell transplantation (alloSCT). In the largest report by Kayser et al. ( Br J Haematol 2021) 21 patients (15 transplanted in first complete remission [CR1], median follow-up 5.5 years) were analyzed. A 5-year overall survival (OS) of 38% was achieved after alloSCT compared to 11% in patients who received chemotherapy only. The aim of our study was to evaluate the outcome of patients with AML harboring t(8;16) after alloSCT in a larger cohort from the EBMT registry. All types of donor, conditioning regimen and graft were included. Sixty patients were identified, of whom 44 had been transplanted in CR1. Median follow-up from alloSCT was 72.3 months, median age was 46.3 years, and 73.3% were female. Karnofsky performance status was &amp;gt;80% in 59 patients, one had missing data. At 2 years after alloSCT, OS and leukemia free survival (LFS) of the entire cohort were 42.7% and 42.2%, respectively, (28.6% and 14.3% in patients transplanted in advanced disease). Cumulative incidence of relapse (RI) and non-relapse mortality (NRM) at two years was 36.9% and 15.3%, respectively, in patients transplanted in CR1, and 78.6% and 7.1%, respectively, in patients transplanted in advanced disease. The 44 patients transplanted in CR1 were analyzed in detail. Basic characteristics were comparable to the entire cohort as described above. Twenty-two of the patients had de novo, 22 had sAML. According to the ELN 2017 classification (not yet including t(8;16) as a factor for poor prognosis), 54.5% had an intermediate and 45.5% an adverse cytogenetic risk. Furthermore, 38.8% had a complex karyotype. Donors were matched related in 34%, matched unrelated in 54%, haploidentical in 5% and cord blood in 7%. Conditioning was myeloablative in 52.3%, whereas 47.7% had received reduced intensity conditioning. Among patients transplanted in CR1, 2- and 5-year OS from alloSCT was 48% and 39.9%, the respective LFS was 46.8% and 35.5%. On multivariate analysis (MVA), harboring t(8;16) within a complex karyotype was the major risk factor for outcome, being associated with higher RI (hazard ratio [HR] 3.38, p=0.01), lower LFS (HR 4.17, p=0.016), and lower OS (HR 3.08, p=0.017). In contrast, patients with t(8;16) outside of a complex karyotype achieved excellent results (Figure 1). Besides a complex karyotype, MVA revealed sAML as an additional, independent risk factor with a higher RI (HR 3.73, p=0.026). Age was the major factor for NRM, reaching 5% vs. 26.5% in patients below or above the median (p=0.02). In conclusion, according to the largest series analyzed in this setting so far, overall results after alloSCT in AML with t(8;16) transplanted in CR1 were favorable, especially in patients without multiple additional cytogenetic abnormalities, achieving a low risk of relapse and a 5-year OS &amp;gt;60%. Hence, alloSCT in CR1 appears to abrogate the unfavorable prognostic value of this translocation. Whether or not t(8;16) does have any additional influence on the outcome of patients with a complex karyotype remains to be evaluated. Figure Legend Figure 1: Outcome of AML with t(8;16) with and without complex karyotype

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 &amp; 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 &amp;lt; 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.

RTK-RAS Signaling Pathway Was Enriched in Rare Acute Myeloid Leukemia Patients with t(16;21)(p11;q22)/ FUS::ERG

Introduction: t(16;21)(p11;q22)/ FUS::ERG is a rare recurrent abnormality in acute myeloid leukemia (AML) and confers a poor prognosis in AML patients. Previous studies analyzed the role of allogeneic hematopoietic stem cell transplantation (HSCT) in these patients but the results were discordant. AML is characterized by concurrent mutations, which cooperated in the leukemogenesis and refractory/relapse mechanism. However, few studies described the mutational landscape of FUS::ERG AML. Olshanskaya et al. discovered concomitant DNMT3A, ASXL1, RUNX1 and BCOR mutations in the FUS::ERG AML patients but the study was limited to the small case number. Therefore, we aim to assess genomic alterations and clinical outcome of FUS::ERG AML patients with a larger cohort. Methods: We reviewed medical records of 12328 AML patients diagnosed between December, 2011 and February, 2022 in our hospital. A total of 39 consecutive AML patients with karyotype or reverse transcription-polymerase chain reaction analysis proven t(16;21)(p11;q22)/ FUS::ERG were identified. Whole exome sequencing was performed in 4 patients and targeted sequencing was performed in 20 patients. Patients with follow-up time longer than 30 days were included in the survival analysis. Overall survival (OS) was calculated from the date of diagnosis to the date of death or censoring. Event free survival (EFS) was calculated from the date of diagnosis to the date of refractory, relapse, death or censoring, whichever came first. Results: M5 was the predominant subtype according to French-American-British classification systems. CD56 (91.9%) and CD123 (81.1%) were found to be expressed in most FUS::ERG AML patients. Six patients had central nervous system involvement diagnosed by flowcytometry (FCM) analysis of cerebrospinal fluid at the time of first lumbar puncture. One patient was diagnosed central nervous system involvement at relapse (Table 1). The most prevalent somatic variants of our FUS::ERG AML patients were PTPN11 (25%), NRAS (20%), RELN (15%), CENPB (10%), RUNX1 (10%) and SH2B3 (10%) mutations (Figure 1). Five patients had no somatic mutation detected by targeted sequencing. Notably, RTK-RAS signaling pathway was the most involved pathway after clustering the aberrated genes according to the Gene Ontology terms, which included PTPN11, NRAS, ABL1 and JAK2 aberrations. All but one PTPN11 mutations affected the amino terminal (N)-SH2 domain. NRAS mutation were found mutated at amino acid G12 and Q61. The median EFS of the patients was 9.3 months (95% CI: 9.2-NA) and OS was 18.2 months (95%CI: 13.1-NA). Due to the poor outcome of this subtype of AML, allogeneic HSCT was highly recommended. A total of 14 patients underwent allogeneic HSCT. Notably, seven of ten patients were positive for minimal residual disease assessed by FCM at the time of transplantation. We did not observe the statistically significant effect of HSCT on improving the EFS (p = 0.24) or OS (p=0.26) of our patients. Conclusions: PTPN11 and NRAS were the most frequent mutations and RTK-RAS signaling pathway was the most involved pathway in rare AML patients with t(16;21)(p11;q22)/ FUS::ERG. The addition of signaling pathway inhibitors followed by HSCT might be an effective strategy to overcome the dismal outcome of this subtype. Larger cohort studies are warranted to investigate the molecular characteristics further as well as evaluate the clinical activity of the SHP2 (PTPN11) inhibitors in FUS::ERG AML patients.

The Co-Occurrence of bZIP in-Frame CEBPA-Mutation with Unfavorable Genetic Abnormalities Is Associated with a Poor Prognosis in Acute Myeloid Leukemia

In a previous study (SY Ahn et al, Cancer Res Treat 2023), we reported that the presence of only the basic leucine zipper in-frame mutation (bZIP in-f) CEBPA is associated with favorable outcomes in patients with CEBPAdouble mutations. However, when bZIP in-fCEBPAmutations were accompanied by certain mutations in chromatic/DNA modifiers, cohesion complex, and splicing genes, patients exhibited inferior overall survival (OS). FLT3-ITD or additional cytogenetic abnormalities is known to rarely co-occur with CEBPA mutation in patients with AML. In cases where both bZIP in-fCEBPA and FLT3-ITD coexist, there are conflicting aspects in risk stratification, and research on its prognostic significance is limited. Therefore, we aimed to evaluate the prognostic significances of the co-occurrence of bZIP in-fCEBPAmutation with unfavorable genetic abnormalities in acute myeloid leukemia. In 832 patients who were diagnosed with AML excluding acute promyelocytic leukemia and underwent intensive induction therapy, 98 (11.8%) had bZIP in-fCEBPA. Among 98 patients with bZIP in-fCEBPA, eight cases had FLT3-ITD (5 with low allelic ratio, 3 with high allelic ratio), 19 patients had unfavorable genetic mutations according to the 2022 European LeukemiaNet guidelines, and 10 patients had non-favorable cytogenetic abnormalities as detected in diagnostic samples. We observed no clinical differences in features such as age, sex, WBC count, bone marrow blast percentage at diagnosis based on the co-occurrence unfavorable genetic abnormalities (unfavorable-gene). Among the 98 patients with bZIP in-f CEBPA, 30 out of 37 patients with unfavorable-gene achieved complete remission (CR) after induction therapy, and 55 out of 61 without unfavorable genetic abnormalities showed CR (p=0.20). Allogeneic hematopoietic cell transplantation was proceeded at first CR in 21 out of 37 unfavorable-gene group, and 35 out of 61 bZIP in-fCEBPAonly group ( p=1.00). In the survival analysis, patients with FLT3-ITD, unfavorable genetic co-mutated, or non-favorable cytogenetic abnormalities with bZIP in-fCEBPA showed inferior OS (p=0.036) (Fig. A). Due to the limited number of patients for analysis, we classified them into two group: the unfavourable-gene group and bZIP in-fCEBPAonly group. We observed that the unfavourable-gene group had a poor OS compared to the bZIP in-fCEBPAonly group (HR 2.71, 95 CI; 1.32-5.55, p=0.007) (Fig.B). On multivariate analysis, unfavorable-gene group was only poorrisk factor for OS. Despite the inclusion of a relatively small number of patients, our study demonstrates that when co-occurrence of bZIP in-fCEBPA withunfavourable genetic abnormalities, it results in a poor prognosis. To confirm its clinical significance, further validation with a larger patient population is necessary.

Single-cell RNA sequencing of a new transgenic t(8;21) preleukemia mouse model reveals regulatory networks promoting leukemic transformation.

T(8;21)(q22;q22), which generates the AML1-ETO fusion oncoprotein, is a common chromosomal abnormality in acute myeloid leukemia (AML) patients. Despite having favorable prognosis, 40% of patients will relapse, highlighting the need for innovative models and application of the newest technologies to study t(8;21) leukemogenesis. Currently, available AML1-ETO mouse models have limited utility for studying the pre-leukemic stage because AML1-ETO produces mild hematopoietic phenotypes and no leukemic transformation. Conversely, overexpression of a truncated variant, AML1-ETO9a (AE9a), promotes fully penetrant leukemia and is too potent for studying pre-leukemic changes. To overcome these limitations, we devised a germline-transmitted Rosa26 locus AE9a knock-in mouse model that moderately overexpressed AE9a and developed leukemia with long latency and low penetrance. We observed pre-leukemic alterations in AE9a mice, including skewing of progenitors towards granulocyte/monocyte lineages and replating of stem and progenitor cells. Next, we performed single-cell RNA sequencing to identify specific cell populations that contribute to these pre-leukemic phenotypes. We discovered a subset of common myeloid progenitors that have heightened granulocyte/monocyte bias in AE9a mice. We also observed dysregulation of key hematopoietic transcription factor target gene networks, blocking cellular differentiation. Finally, we identified Sox4 activation as a potential contributor to stem cell self-renewal during the pre-leukemic stage.

A novel t (5; 17) (q35; q21) associated with t (8; 21) (q22; q22) in a patient with acute myeloid leukemia: case report and review of literature.

The t (8; 21) (q22; q22) with the resulting RUNX1- RUNX1T1 rearrangement is one of the most common cytogenetic abnormalities in acute myeloid leukemia (AML). It is associated with favorable prognosis. The t (5; 17) (q35; q21) is an uncommon translocation, fuses the gene for the nucleophosmin (NPM) to the retinoic acid receptor α(RARA) and was described essentially in acute promyelocytic leukemia (APL) variant. We present the case of a 19-year-old male patient who developed an AML with t (8; 21) (q22; q22) associated to t (5; 17) (q35; 21). Morphology and immunophenotype of the leukemic cells were compatible with AML. The patient received chemotherapy based on cytarabine and anthracycline without all-trans retinoic acid (ATRA) followed by allogenic stem cell transplantation in first remission. To the best of our knowledge, this is the first report of an association between a rare translocation t (5; 17) and t (8; 21) in AML. In this report, we will discuss the prognosis of this association as well as the treatment.

Outcomes of Patients with FLT3 Positive Acute Myeloid Leukaemia; an Experience from a Tertiary Care Hospital in Karachi, Pakistan.

Molecular genetic abnormalities in acute myeloid leukaemia (AML) are essential for disease diagnosis and determining prognosis and clinical course. Mutations in FLT3 and nucleophosmin (NPM) genes are the most frequent genetic abnormalities, which are also known to impact disease outcomes. FLT3 mutations have been identified in approximately 30% of de novo AML patients and are associated with poor prognoses. This study aimed to determine the response to induction chemotherapy, overall survival (OS) and relapse rate (RR) in patients with FLT3-positive AML. In this study, a retrospective analysis was performed of 75 newly diagnosed patients with AML registered between January 2015 and July 2022. Patient demographics and clinical-haematological parameters were noted and molecular analysis for FLT3 ITD/TKD and NPM mutations was performed. All the patients received standard induction chemotherapy and their response to treatment, OS and RR were assessed. A total of 75 cases of AML were analysed. The mean age of the sample was 34.9 years, of which 65.3% were males and 34.7% were females. The patients were stratified into two groups: Those who were positive for FLT3 while negative for NPM (FLT3+/NPM-), representing 17.3% and those who were negative for both FLT3 and NPM (FLT3-/NPM-), representing 82.7% of cases. On day 28 post-induction, the complete remission rate was 69.2% in the FLT3 positive group and 77.4% in the FLT3 negative group. In the FLT3+/NPM- group, 55.6% of cases who were in remission at day 28 subsequently relapsed, compared to 50.0% of FLT3-/NPM- cases. The overall median survival time for the cohort and FLT3+ group was 1467 days, while that of the FLT3-group could not be estimated due to the very high survival rate. No significant differences in outcomes were observed in patients who were FLT3 positive compared to those who were FLT3 negative.

Role of Gene Mutations in Acute Myeloid Leukemia: A Review Article

AbstractAcute myeloid leukemia (AML) is an immensely heterogeneous disease characterized by the clonal growth of promyelocytes or myeloblasts in bone marrow as well as in peripheral blood or tissue.Enhancement in the knowledge of the molecular biology of cancer and recognition of intermittent mutations in AML contribute to favorable circumstances to establish targeted therapies and enhance the clinical outcome. There is high interest in the development of therapies that target definitive abnormalities in AML while eradicating leukemia-initiating cells. In recent years, there has been a better knowledge of the molecular abnormalities that lead to the progression of AML, and the application of new methods in molecular biology techniques has increased that facilitating the advancement of investigational drugs.In this review, literature or information on various gene mutations for AML is discussed. English language articles were scrutinized in plentiful directories or databases like PubMed, Science Direct, Web of Sciences, Google Scholar, and Scopus. The important keywords used for searching databases is “Acute myeloid leukemia”, “Gene mutation in Acute myeloid leukemia”, “Genetic alteration in Acute myeloid leukemia,” and “Genetic abnormalities in Acute myeloid leukemia.”

Molecular and cytogenetics abnormalities in acute myeloid leukemia at Puerto Rico.

e19005 Background: Acute myeloid leukemia (AML) is a highly heterogeneous disease. Specific cytogenetic and molecular features permit stratification of this disease. Prognosis within these categories varies widely. Determining optimal management has been complicated by the multiple molecular disease subsets with different responses to standard therapeutics. The comprehensive genetic analysis of AML in the past 10 years has revealed recurrent, somatically mutated genes, leading to the development of new targeted therapies including inhibitors of FLT3 and IDH. Decisions about the choice of therapy in patients with AML currently depend on the identification of a selected set of genetic markers at diagnosis. At Puerto Rico, AML data of cytogenetic and molecular markers is lacking. According to the Puerto Rico Cancer Registry from 2015 to 2019 a total of 551 cases of AML were reported. The San Juan City Hospital-hematology oncology department is one of the major referral centers on the island for initial evaluation and management of AML. The objective of our retrospective study was to identify the most common cytogenetic and molecular abnormalities present in our population. Patient conditions were stratified as favorable, intermediate, or high risk, in terms of prognosis using the ELN revised 2022 stratification. Methods: This study examined the frequency of molecular markers in newly diagnosed (ND) patients with AML. We evaluated patients that were at least 18 years of age or older with cytogenetic and molecular analysis in a period of 4 years (2019-2022). All data regarding demographic, bone marrow biopsy, cytogenetics, and molecular markers were obtained from electronic medical record at our institution with institutional review board approval. Results: A total of 140 patients with ND AML were identified during the study period. 58% female and 42% male with a median age of 57 years old at time of diagnosis. In terms of molecular markers, the most common were RUNX1 (22%), follow by IDH2 (14%), FLT3-ITD (14%) and TP53 (11%). Complex cytogenetics was identified in 40% patients, and 34% had Myelodysplasia related cytogenetic. Using the ELN 2022 criteria to classify AML patients in terms of risk with cytogenetics and molecular markers, our study revealed 21% favorable risk, 31% intermediate, and 48% adverse. Conclusions: Our study suggests that our population has higher prevalence of AML with myelodysplasia-related gene mutations and molecular related to adverse risks, such as RUNX1. Further studies with appropriate sub-group analysis need to be followed to determine if there is any genetical predisposition on the puertorrican population for myelodysplasia related genes or for RUNX1.

Whole transcriptome sequencing reveals HOXD11-AGAP3, a novel fusion transcript in the Indian acute leukemia cohort.

Introduction: Acute leukemia is a heterogeneous disease with distinct genotypes and complex karyotypes leading to abnormal proliferation of hematopoietic cells. According to GLOBOCAN reports, Asia accounts for 48.6% of leukemia cases, and India reports ~10.2% of all leukemia cases worldwide. Previous studies have shown that the genetic landscape of AML in India is significantly different from that in the western population by WES. Methods: We have sequenced and analyzed 9 acute myeloid leukemia (AML) transcriptome samples in the present study. We performed fusion detection in all the samples and categorized the patients based on cytogenetic abnormalities, followed by a differential expression analysis and WGCNA analysis. Finally, Immune profiles were obtained using CIBERSORTx. Results: We found a novel fusion HOXD11-AGAP3 in 3 patients, BCR-ABL1 in 4, and KMT2A-MLLT3 in one patient. Categorizing the patients based on their cytogenetic abnormalities and performing a differential expression analysis, followed by WGCNA analysis, we observed that in the HOXD11-AGAP3 group, correlated co-expression modules were enriched with genes from pathways like Neutrophil degranulation, Innate Immune system, ECM degradation, and GTP hydrolysis. Additionally, we obtained HOXD11-AGAP3-specific overexpression of chemokines CCL28 and DOCK2. Immune profiling using CIBRSORTx revealed differences in the immune profiles across all the samples. We also observed HOXD11-AGAP3-specific elevated expression of lincRNA HOTAIRM1 and its interacting partner HOXA2. Discussion: The findings highlight population-specific HOXD11-AGAP3, a novel cytogenetic abnormality in AML. The fusion led to alterations in immune system represented by CCL28 and DOCK2 over-expression. Interestingly, in AML, CCL28 is known prognostic marker. Additionally, non-coding signatures (HOTAIRM1) were observed specific to the HOXD11-AGAP3 fusion transcript which are known to be implicated in AML.

Somatic exonic deletions in RUNX1 constitutes a novel recurrent genomic abnormality in acute myeloid leukemia.

In acute myeloid leukemia (AML), somatic mutations (commonly missense, nonsense and frameshift indels) in RUNX1 are associated with a dismal clinical outcome. Inherited RUNX1 mutations cause familial platelet disorder. As approximately 5-10% of germline RUNX1 mutations are large exonic deletions, we hypothesized that such exonic RUNX1 aberrations may also be acquired during the development of AML. Sixty well-characterized AML patients were analyzed with Multiplex Ligation-dependent Probe Amplification (MLPA, n=60), micro-array (n=11) and/or whole genome sequencing (WGS, n=8). In total, 25 (42% of the cohort) RUNX1-aberrant patients (defined by the presence of classical mutations and/or exonic deletions) were identified. Sixteen patients (27%) carried only exonic deletions, 5 (8%) carried classical mutations and 4 (7%) carried both exonic deletions and mutations. No significant difference was observed between patients with classical RUNX1 mutations and RUNX1 exonic deletions in median overall survival (OS, 53.1 vs 38.8 months respectively, p=0.63). When applying the European Leukemia Net (ELN) classification including the RUNX1-aberrant group, 20% of the patients initially stratified as intermediate-risk (5% of the whole cohort) were re-assigned to the high-risk group, which improved the performance of ELN classification regarding OS between intermediate and high-risk group (18.9 vs 9.6 months, p=0.09). Somatic RUNX1 exonic deletions constitute a novel recurrent aberration in AML. Our findings have important clinical implications regarding AML classification, risk-stratification and treatment decision. Moreover, they argue in favor of further investigating such genomic aberrations not only in RUNX1 but also in other genes implicated in cancer biology and management.

Targeted Therapy Development in Acute Myeloid Leukemia

Therapeutic developments targeting acute myeloid leukemia (AML) have been in the pipeline for five decades and have recently resulted in the approval of multiple targeted therapies. However, there remains an unmet need for molecular treatments that can deliver long-term remissions and cure for this heterogeneous disease. Previously, a wide range of small molecule drugs were developed to target sub-types of AML, mainly in the relapsed and refractory setting; however, drug resistance has derailed the long-term efficacy of these as monotherapies. Recently, the small molecule venetoclax was introduced in combination with azacitidine, which has improved the response rates and the overall survival in older adults with AML compared to those of chemotherapy. However, this regimen is still limited by cytotoxicity and is not curative. Therefore, there is high demand for therapies that target specific abnormalities in AML while sparing normal cells and eliminating leukemia-initiating cells. Despite this, the urgent need to develop these therapies has been hampered by the complexities of this heterogeneous disease, spurring the development of innovative therapies that target different mechanisms of leukemogenesis. This review comprehensively addresses the development of novel targeted therapies and the translational perspective for acute myeloid leukemia, including the development of selective and non-selective drugs.

Novel high–risk acute myeloid leukemia subgroup with ERG amplification and Biallelic loss of TP53

ETS-related gene (ERG) amplification, observed in 4–6% of acute myeloid leukemia (AML), is associated with unfavorable prognosis. To determine coincident effects of additional genomic abnormalities in AML with ERG amplification (ERGamp), we examined 11 ERGamp cases of 205 newly diagnosed AML using chromosomal microarray analysis and next generation sequencing. ERGamp cases demonstrated a distinct pattern of high genetic complexity: loss of 5q, chromothripsis and TP53 loss of function variants. Remarkably, allelic TP53 loss or loss of heterozygosity (LOH) co-occurring with TP53 inactivating mutation dramatically effected ERGamp tumor patient outcome. In the presence of homozygous TP53 loss of function, ERGamp patients demonstrated no response to induction chemotherapy with median overall survival (OS) of 3.8 months (N = 9). Two patients with heterozygous loss of TP53 function underwent alloSCT without evidence of relapse at one year. Similarly, a validation TCGA cohort, 6 of the 8 ERGamp cases with TP53 loss of function demonstrated median OS of 2.5 months. This suggests that with TP53 mutant ERGamp AML, successive loss of the second TP53 allele, typically by 17p deletion or LOH identifies a specific high-risk subtype of AML patients who are resistant to standard induction chemotherapy and need novel approaches to avert the very poor prognosis.

Frequency and pattern of chromosomal abnormalities in acute myeloid leukemia from Western India: A retrospective study

Chromosomal abnormalities play an important role in diagnosis and prognosis of hematological diseases. The aim of the present study was to study the pattern and frequency of chromosomal aberrations in acute myeloid leukemia (AML) subgroups from western India. A retrospective study was conducted through evaluating laboratory proforma which were filled during 2005 to 2014 for diagnosis and treatment of AML subjects. We have studied chromosomal aberrations in 282 subjects with AML from western India. AML patients were sub-grouped according to FAB classification. Cytogenetic study using conventional cytogenetics (GTG-banding) and Fluorescence in situ hybridization (FISH) was carried out using FISH probes (AML1/ETO, PML/RARA, CBFB). Student's t test for continuous variables and Pearson's Chi-squared test for categorical variables were used to identify the relationship between variables. Cytomorphological study revealed AML- M3 as most frequent (32.3%) group followed by AML-M2 (25.2%) and AML-M4 (19.9%). Chromosomal abnormalities were identified in 145 (51.42%) of the total AML cases. A high frequency (38.6%) of chromosomal abnormalities was identified in AML-M3 subgroup as compared to AML-M2 (31%) and AML-M4 (20.6%). Cytogenetic study is important for the diagnosis and management of the AML patients. Our study identified chromosomal abnormalities in AML subgroups with varied frequencies. It is important in diagnosis and monitoring of the disease. As younger AML patients were more affected in our study, etiological factors such as environmental factors need to be studied. Combination of conventional cytogenetics and FISH has an advantage of identifying high frequency of chromosomal aberrations in AML patients.