Related Acute Myeloid Leukemia Research Articles

Prognostic Value of Copy Number Aberrations and Copy Neutral Loss of Heterozygosity in Acute Myeloid Leukemia

Background: Chromosomal abnormalities are key to the diagnosis and prognosis of patients with acute myeloid leukemia (AML). Chromosomal genomic array testing (CGAT) can identify copy neutral loss of heterozygosity (cnLOH) and genomic copy number alterations (CNAs) that are undetected by conventional karyotype or fluorescence in situ hybridization (FISH). Previous studies have shown that cnLOH is correlated with a shorter duration of complete remission (CR) and worse overall survival (OS) in AML (Grosneth et. al, Cancer, 2015). The primary objective of this exploratory study is to examine the predictive power of CGAT in a cohort of AML patients with majority normal karyotype. Methods: CGAT analyses were collected from 187 subjects with AML or other high grade myeloid neoplasm (myeloid blasts >10% in peripheral blood or bone marrow) between 2012 and 2022 at Fred Hutchinson Cancer Center. CGAT utilized CytoScan™ HD Array (ThermoFisher, MA), targeting genome-wide region with 2.4 million markers for copy number and approximately 750,000 genotype-able SNPs. The data were analyzed using the Nexus Copy Number software with manual curation. Based on the assay validation in our CLIA-certified diagnostic lab, the cutoff is 100 Kb for copy number aberrations and 10 Mb for cnLOH. Survival predictive power analysis (SPP), a log-rank based test using Nexus, was applied to identify aberrations significant for event free survival (EFS) and overall survival (OS). SPP analysis tabulated genomic segments based on p-values with the alpha value cutoff at 0.05. Using frequency as the filtering criteria (present in >5 patients), 132 SPP for EFS, and 73 SPP for OS were identified. SPP were then manually curated to (1) remove those representing constitutional absence of heterozygosity (AOH) and those overlapping with benign copy number variance and (2) connect adjacent SPP into contiguous regions. The presence and absence of the identified SPP were then tabulated, cleaned up, and used to generate Kaplan-Meier curves for EFS and OS. This process identified 5 SPP that are significant for EFS and 2 SPP for OS. A multivariate analysis was performed to determine the hazard ratio (HR) of these 5 genetic aberrations for EFS and OS, adjusted for patient age, secondary/treatment related AML, ELN 2022 risk, and receipt of allogeneic hematopoietic cell transplantation (HCT) as a time-varying covariate. Results: In our 187 subject cohort, the median age was 63 (range 20-87). 76 (41%) had secondary or treatment related disease. 97 (52%) went on to receive a hematopoietic stem cell transplant. 3 had favorable risk disease, 14 had adverse risk, and 170 had intermediate risk by karyotype and FISH using ELN 2022 criteria. 120 (64%) had normal karyotype. 5 subjects had 4q cnLOH, 6 had 5q ( EGR1) loss, 7 had 9p cnLOH, 10 had mixed-lineage leukemia gene partial tandem duplication ( MLL-PTD), and 4 had 7q deletion. Univariate analysis identified five SPP that are significant for EFS: 4q cnLOH (p=0.002), 5q ( EGR1) loss (p<0.001), 9p cnLOH (p<0.001), MLL-PTD (p<0.001), and 7q deletion (p<0.001). 2 SPP were significant for OS: 9p cnLOH (p=0.001) and MLL-PTD (p=0.036). Multivariate analysis for EFS adjusted for age, secondary/treatment related disease, ELN 2022 risk, and receipt of HCT as a time-varying covariate demonstrated a hazard ratio (HR) of 3.5 for 4q cnLOH (p=0.008), 2.3 for 5q ( EGR1) loss (p=0.16), 4.5 for 7q deletion (p=0.007), 5.7 for 9p cnLOH (p<0.001), and 2.9 for MLL-PTD (p=0.002). Multivariate analysis for OS adjusted for age, secondary/treatment related disease, ELN 2022 risk, and receipt of HCT as a time-varying covariate demonstrated a hazard ratio (HR) of 2.0 for 4q cnLOH (p=0.23), 0.5 for 5q ( EGR1) loss (p=0.36), 1.3 for 7q deletion (p=0.66), 4.9 for 9p cnLOH (p=0.001), and 2.3 for MLL-PTD (p=0.04). Discussion: This study has found 5 key genetic changes identifiable via CGAT that are significant for shorter EFS and/or OS in AML patients. Our next step is to validate these findings in a large independent cohort of AML patients. CGAT is a powerful adjunct to conventional karyotype and FISH for the detection of cnLOH and can be used to identify CNAs below the level of detection of karyotype. For these reasons, CGAT may be most useful in intermediate risk patients. Further research is warranted to fully understand the prognostic power of CGAT in AML.

Burden of Chemotherapy-Related Myelodysplastic Syndrome/Acute Myeloid Leukemia in the Chemotherapeutic Armamentarium

Introduction: Therapy related Myelodysplastic Syndrome and Acute Myeloid Leukemia (t-MDS/AML) is a well-established adverse event (AE) associated with conventional chemotherapy drugs and has historically carried a poorer prognosis when compared to de novo disease. Newer chemo medicines like PARP inhibitors have also been implicated in developing this complication. However, comprehensive real-world data is lacking pertaining to the AE burden relative to individual drug classes. Methods: We used the FDA Adverse Events Report System (FAERS) public dashboard to obtain the data. Data was available for the years 1970 - 2023. We retrieved the number of events that were reported as “Myelodysplastic Syndrome” or “Acute Myeloid Leukemia” for individual chemotherapy classes that have historically been known to cause t-MDS/AML. Disproportionality analysis was performed by calculating the reporting odds ratio (ROR) with its 95% confidence interval (CI). Results: The drug class with the highest ROR of MDS/AML was the topoisomerase inhibitors - ROR 2.38 (95% CI 2.27-2.49). Intercalating agents like anthracyclines and mitoxantrone had the next highest ROR of 2.12 (95% CI 2.04 - 2.21). The novel drug class PARP inhibitors had the third most burden of t-MDS/AML with an ROR of 1.57 (95% CI 1.46 - 1.69). Table 1. depicts the individual drug classes and their ROR for associated MDS/AML. Table 1 depicts the individual drug classes and their ROR for associated MDS/AML. Conclusions: From this retrospective analysis, we identified that among all the chemotherapy drug classes, the topoisomerase II inhibitors were associated with the highest burden of t-MDS/AML, followed by intercalating agents and PARP inhibitors. Further data is essential to identify the patient characteristics associated with higher risk of developing this dreaded complication. This could improve patient-counseling prior to treatment initiation, assist with alternate drug choices when applicable, and perhaps also aid in developing strategies for closer monitoring in high-risk patients.

Identification, Monitoring and Reversibility of PARP Inhibitor Associated Clonal Haematopoiesis and Myelodysplastic Syndrome: A Case Series

Background: Poly ADP ribose polymerase inhibitors (PARPi) are widely used as maintenance treatment for ovarian cancer (OC) after response to platinum chemotherapy in the first line setting and recurrent disease. Their use has significantly improved outcomes, particularly in patients with BRCA mutations in whom first-line PARPi maintenance therapy increases progression-free survival from 13.8 to 56.0 months (Hazard Ratio, 0.33; 95% CI, 0.25-0.43). However, a recent meta-analysis of clinical trial adverse events and pharmacovigilance data showed that PARPi therapy significantly increases the risk of therapy related myeloid neoplasms (tMN), myelodysplastic syndrome (MDS) and acute myeloid leukaemia with poor outcomes. Platinum based chemotherapy is the mainstay of OC treatment and is associated with clonal haematopoiesis (CH) with mutations in DNA damage response genes. CH is recognised as a premalignant myeloid state but real world data on clonal dynamics during PARPi therapy are lacking. We present a case series of OC patients receiving PARPi-therapy referred for haematological review for beyond-expected HT or unexplained full blood count (FBC) abnormalities with serial FBC and myeloid next generation sequencing (mNGS) clonal monitoring in relation to PARPi therapy. Methods: Seven patients with stage III/IV high-grade OC were referred to haematology at the discretion of the treating oncologist between February 2017 and May 2022 (mean age (SD): 68 (7.0) years, median PARPi duration: 18 months (range: 4-86)). All patients had received prior platinum-based chemotherapy and were screened with mNGS in peripheral blood (PB) or bone marrow aspirate and trephine (BMAT) at initial assessment. Results: Reasons for referral are detailed in Table 1 and include two patients with isolated elevated nucleated red blood cell (NRBC) counts. Six patients had clonal pathology (CP), five of whom had single-nucleotide variants typical of platinum-exposure defined by the involved gene or platinum-associated mutational signatures. Clinical details including BRCA-status, haematological diagnosis and mNGS results are detailed in Table 1. PARPi-therapy was discontinued in four patients with CP, with resolution or improvement of FBC and m-NGS abnormalities (variant allele frequencies (VAFs) and/or copy number variations (CNVs)) in three patients. These findings were most marked in Patient 1, referred with grade 3 thrombocytopenia and transfusion dependence and in whom PARPi cessation resulted in transfusion independence and resolution of morphological and cytogenetic abnormalities (clinical course including BMAT results and haematological parameters illustrated in Figure 1), and Patient 3, who demonstrated selective regression of a TP53 mutated clone on PARPi cessation. Two patients with germline BRCA mutations continued PARPi therapy with 3 monthly PB mNGS monitoring after discussion at multidisciplinary team meetings. Conclusions: We show using sequential molecular and haematological monitoring before and after PARPi cessation that morphological, FBC and clonal abnormalities can be reversed on PARPi discontinuation, potentially mitigating the risk of life-threatening tMNs in some patients. We also highlight that, in addition to cytopenias, persistently elevated NRBCs may be an indicator of clinically relevant CP not well described in pharmacovigilance data based on common terminology criteria for adverse events criteria. Large scale prospective data on clonal dynamics in PARPi therapy are needed to inform mNGS screening and tMN risk stratification as scoring systems based on age associated clonal haematopoiesis of indeterminate potential may not be applicable. Until these data are available we demonstrate the value in MDT discussion and serial PB mNGS monitoring in formulating case-by-case therapeutic decisions regarding CP and PARPi therapy, particularly patients with high risk BRCA mutated OC.

Significance of the Apoptotic Regulators BAX, BCL2, BCL- XL and MCL1 in Newly Diagnosed AML

Introduction: In acute myeloid leukemia (AML), cytogenetics and molecular mutations remain important prognostic tools for tailoring treatment after induction therapy. Abnormalities in multiple components of apoptotic pathways have been identified as potential drivers of AML. Identifying other biomarkers may help us further refine prognosis and identify other therapeutic strategies. B-cell lymphoma 2 (Bcl2), Bcl-xL and myeloid cell leukemia-1 (Mcl1) are commonly expressed prosurvival proteins in hematologic malignancies, whereas Bax acts as a promoter of apoptosis. We retrospectively evaluated the significance of Bax, Bcl2, Bcl-xL and Mcl1 expression in newly diagnosed AML patients using the Beat AML program 1. Methods: We analyzed RNA-expression data from The Beat AML study (672 tumor specimens collected from 562 patients) 1. Data was extracted from the public database. Different expression levels of the above genes were compared using t-tests. Results were considered significant with p-value < .05. Kaplan-Meier curves were used to estimate survival. Results: For the correlative survival analyses, Bax, Bcl2, Bcl-xL and Mcl1 mRNA expression and clinical information were both obtained from the BeatAML cohort 1. Patients were categorized into high expressers (≥ median) and low expressers (<median). In total, 461 patients had available gene expression data. The mean age of diagnosis was 57 years, and 56.6% of patients were male. The most common specific diagnosis at inclusion was AML with mutated NPM1 (22%), followed by AML with myelodysplasia related changes (21%) and AML, NOS (11%). Complete response to induction was seen in more than half (56%) of the patients for which response was recorded (n=369). Others developed refractory AML (32%). No data was available for 12%. Standard chemotherapy (induction therapy with idarubicin and cytarabine (7+3) and consolidation therapy with high dose cytarabine) was used in 98.4% of these patients. Over half of the analyzed samples were peripheral blood (52%), and the remaining samples were either bone marrow aspirate (46%) or from leukapheresis (2%). Most of the specimens were collected at initial acute leukemia diagnosis (62.2%), others were collected with residual disease or post chemotherapy (25.1%). The mean survival for this cohort was 13.7 months. Above median expression of Mcl1 was associated with longer survival (33 vs. 20 months; p = 0.0262). Above median expression of Bax was also associated with longer survival (32 vs. 18.2 months; p = 0.0114). In the Bax analysis, gene expression was further subdivided into 4 quartiles which showed correlation of expression levels with length of survival (Figure 1). The prognostic significance of the ratio Bax/ (Mcl1+Bcl2+Bcl-xL) was also analyzed given the mechanism of action of these various regulators. A higher ratio was associated with longer survival (40.7 vs. 18.7 months; p = 0.0095) (Figure 2) Conclusions Bcl-2, Bcl-xL and Mcl-1 are commonly expressed antiapoptotic proteins, whereas Bax is a promoter of apoptosis. The results with Bax are consistent with previous literature. Our results with Mcl1 expression differed from what we may have expected. This may be due to the sample size or methodology. However, it may also be due to the fact that a ratio of the actors is needed to help predict prognosis (as we analyzed in Figure 2). The overall survival between Bcl-2 and Bcl-xL was compared independently, however, they did not appear to have a significant difference in survival between high and low expression groups. We are currently working to see if this ratio can be validated in a prospective cohort of relapsed/ refractory AML patients treated on clinical trial and this data will be presented at the meeting. The Bcl-2 inhibitor venetoclax has demonstrated promising results in treating AML. However, venetoclax appears to have limited activity as a single agent. Our study suggests that potentially a combination of agents targeting factors such as Bcl-2, Bcl-xL and Mcl-1 may be associated with improved survival in patients with AML. Further validation of these biomarkers is warranted, and Mcl-1 inhibitors are currently in clinical trials.

Venetoclax Combined with Azacitidine and Homoharringtonine in Adults with Secondary Acute Myeloid Leukemia

Secondary acute myeloid leukemia is an aggressive subset of acute myeloid leukemia (AML) that is, in general, is associated with a very poor prognosis and high mortality. Venetoclax combined with hypomethylating agents have showed relatively successful therapeutic effective in patients with relapse/refractory AML, while efficacy is limited in high-risk subset AML. We hypothesised that Venetoclax combined with Azacitidine and Homoharringtonine could have improved activity in patients with secondary acute myeloid leukemia, particularly in high-risk subgroups. Methods This single center, phase 2 trial was done at the first affiliated hospital of Nanjing medical university. The study enrolled adult patients with diagnosed of secondary AML (AML progressed after preexisting myeloid neoplasms or treatment AML arising from prior exposure of leukemogenic therapies). Patients were required to have an Easter Cooperative Oncology Group performance status of 3 or less, and adequate end-organ function. Patients who had received previous BLC2 inhibitor therapy were excluded. Patients received Azacytidine 75mg/m 2 and Homoharringtonine 3mg for 7 days with venetoclax 400mg daily. The primary endpoint was overall response rate. The secondary endpoints include safety, overall survival, and duration of response (according to recommendations of European LeukemiaNet 2017 guidelines). The trial was registered on ClinicalTrial.gov (NCT 05513131) and continues to accrue patients. Results Since now, we enrolled 11patients; 8(72.7%) patients had secondary AML, 3 (27.3%) patients had treatment related AML, while 2(18.2%) patients had TP53 mutation. The median age was 62 years (48-78 years). At the July 30, 2023 data cutoff, median follow-up time was 36.7 weeks (range 6.57-82.29 weeks). The overall response rate was 81.8% (9 of 11 patients). The most common treatment-emergent adverse events included bone marrow suppression with grades 3 or 4 neutropenia (n=11/11) and febrile neutropenia (n=10/11). The 30-day mortality for all patients was 9.1% (n=1). 2(18.2%) patients were eligible for allogeneic hematopoietic stem cell transplantation. The median overall survival was not achieved. Conclusion Venetoclax combined with Azacitidine and Homoharringtonine regimen had a tolerable safety profile and showed encouraging clinical activity characterised by a high response rate. further patients are needed to validate these findings.

Socioeconomic Status Is a Mediator of Overall Survival in Acute Myeloid Leukemia (AML)

Introduction: AML is a morbid disease with a five-year overall survival (OS) rate of 30.5%. Despite increasing treatment options and improved OS, inequities exist. Some studies cite a 3-year OS difference between Black and White AML patients (pts) at 34% versus 43% (p < 0.001) (Bhatnagar 2021). This study examines data from AML pts in community and academic centers across the United States to assess for differences in outcomes by demographic factors. Methods: This study used the nationwide Flatiron Health electronic health record derived de-identified database with a data cutoff date of June 30, 2022. Pts were over 18 years old with a diagnosis of AML were included. Pts with acute promyelocytic leukemia or therapy related AML were excluded. The primary outcome was OS defined as time between date of diagnosis and date of death or last follow up stratified by demographic factors of race, ethnicity, sex, and socioeconomic status (SES). Area-level SES is an index quintile based on an area-level measure providing insight into pts' neighborhood SES conditions with a score of 1 representing the lowest SES and 5 representing the highest (Guadamuz 2022). Secondary outcomes included first line (1L) treatment stratified by demographic factors. Results: 3338 pts were included in the analysis. 59.8% of pts were age 65 or older, 55.9% were male, and 93.5% were non-Hispanic. 75.2% reported as White, 7.9% as Black or African American, 14.7% as other and 2.1% as Asian. Table 1 highlights key demographic features of the pts with European Leukemia Net (ELN) risk stratification. With a median follow up of 3.33 years, the median OS for the population was 15.9 months (mos) (95% CI 15, 16.9). When broken down by race, median OS was 21.4mos for Asian pts (95% CI 12.6, 23), 18.4mos for Black pts (95% CI 15, 24), 15.4mos for other pts (95% CI 13.6, 18.2), and 16.1mos for White pts (95% CI 14.9, 17.1). These results were not statistically significant (p = 0.67). When stratified by age group, there was a marginally significant difference in OS with Black pts showing a decreased OS in the age group 18-40 years. Median OS for Hispanic pts was higher at 22.7mos (95% CI 17.4, 35.9) compared to non-Hispanic pts at 15.3mos (95% CI 14.4, 16.4) (p = 0.0010). Female sex was associated with improved OS at 17.7mos (95% CI 15.6, 19.3) versus 15mos for male (95% CI 13.8, 16.1) (p < 0.0001). Median OS by SES had improved OS for pts with SES status 4 or 5 at 17.2mos (95% CI 15.7, 18.6) compared to pts with SES 1 or 2 at 16.1mos (95% CI 14.1, 18.0) (p = 0.03) (Fig.1). The mediating effect of SES on race was explored using Cox regression analysis, multivariate models, mediation analysis, and multivariate logistic regression. Analysis indicated no significant mediating effect (p = 0.3142, p = 0.1377, p = 0.3929 for cde/nde, nie, and total effect respectively). 1L treatment analysis showed greater use of 7+3 in Black versus White pts (25% vs 21%) and greater use of hypomethylating agent (HMA) or HMA+venetoclax in White versus Black pts. Similar findings were seen in the Hispanic versus Non-Hispanic pts, with higher use of 7+3 in Hispanic pts (32.5% vs 21.5%). These findings will be further stratified by age group and ELN risk category. There were no significant differences in the use of 7+3, HMA or HMA+venetoclax based on SES category. Discussion: This is a population-based study of AML pts in academic and community centers. Prior studies demonstrated Black pts have worse OS versus White pts. Similarly, we saw a non-statistically significant trend of decreased OS in young Black pts compared to White pts, but no difference when comparing races when assessing patients with an age cut off of 65 years. Hispanic White pts had statistically significant improved OS versus non-Hispanic pts, a finding not noted previously. It is possible with novel regimens such as HMA/venetoclax are responsible for improved outcomes, particularly in the elderly population, although this needs to be further explored. Molecular analysis could yield additional information on the effect of certain mutations and OS. Abraham et al (2022) demonstrated structural racism played a significant role in AML treatment outcomes in pts treated at Chicago academic centers. In our study, lower SES had a negative impact on OS, even when accounting for other factors. These findings are similar to a study by Rebechi et al (2023) and reveal the need for prospective research to mitigate structural barriers to care for AML pts.

CLEC2A Is a Novel AML-Restricted Immunotherapeutic Target Enriched in KMT2A-Rearranged Acute Myeloid Leukemia

KMT2A rearrangements (KMT2A-r) are a frequent event in childhood acute myeloid leukemia (AML) and in treatment related AML in adults leading to a highly refractory and deadly subtype of AML regardless of age. Targeted therapies for KMT2A-r AML have had limited success with novel therapies urgently needed. Despite successes in other leukemias, immunotherapy in AML is in its infancy in part due to significant overlap of antigens expressed in AML and normal hematopoietic cells, which would limit efficacy and increase hematopoietic toxicity. We interrogated the AML transcriptome compared to normal hematopoiesis to identify optimal targets that are silent in normal hematopoieisis and are highly expressed in AML. Here we describe a novel immunotherapeutic target, CLEC2A, a member of the C-type lectin family, that is highly enriched in high-risk AML, particularly KMT2A-r AML and absent in normal hematopoeisis. We evaluated the association between KMT2A fusions and CLEC2A expression identifying CLEC2A as a putative oncoprotein and the potential for CLEC2A to serve as an immunotherapeutic target in high-risk AML via antibody-mediated cytotoxicity. Diagnostic specimens from 1,864 patients with AML (Pediatric 0-29 years from COG Phase III trials N=1486; Adult 18-88 years SWOG N=199 and TCGA-LAML N=179) were interrogated using contemporary NGS to identify genetic aberrations. Conventional karyotyping and mutational analysis were used to determine additional cytogenetic and molecular abnormalities. As CLEC2A was entirely absent in normal bone marrow (TPM 0-0.15), positive CLEC2A expression was defined as mRNA transcripts per million (TPM) >1. Although, CLEC2A was detected in only 18% of AML patients (252/1486; 17% pediatric and 90/378; 24% adult patients), it was highly enriched in those with KMT2A-r AML with 77% (N=193) of CLEC2A positive patients having KMT2A-r, accounting for 48% of the total KMT2A-r cohort, Fig 1A. CLEC2A expression was enriched in specific fusion subgroups: MLLT10 (80%), MLLT4 (71%), and MLLT3 (48%), Fig 1B. CLEC2A expression was low to absent in a majority of other patients, though was seen in other high-risk groups including non-KMT2A MLLT10 fusions, NUP98 fusions, CBFA2T3-GLIS2 fusions, and monosomy 7. CLEC2A is also expressed in a subtype of T-ALL (HOXA activated), which is a fusion driven T-ALL. We further evaluated a potential causal link between KMT2A fusions and CLEC2A expression and inquired whether KMT2A fusion proteins might directly bind to the CLEC2A promoter and induce CLEC2A expression. We used AutoCUT&RUN profiling of genome-wide occupancy of KMT2A oncoproteins to evaluate direct binding of the KMT2A fusion protein to the CLEC2A promoter in primary AML patient samples with three different KMT2A fusions (MLLT10, MLLT4 and MLLT3). We found direct binding of KMT2A fusion proteins to the CLEC2A promoter region, indicating a causal relationship between KMT2A fusions and CLEC2A expression. We then evaluated the potential of therapeutic targeting of CLEC2A by an antibody-drug conjugate using a HumZap cytotoxicy assay. CLEC2A antibody conjugated to saporin (a toxin with cytolytic activity when internalized) was incubated with CLEC2A+ and CLEC2A- cells with target-specific cytotoxicity evaluated by colorimetric assay. CLEC2A+ OCI-AML2 cells had significantly higher rate of cytotoxicity compared to CLEC2A- control cells at 10nM Ab-toxin conjugate. Clinical outcomes for patients with and without CLEC2A expression was evaluated. Patients with CLEC2A had significantly worse outcomes (EFS 23% vs. 45.5%, p<0.0001 and OS 38% vs. 63%, p<0.0001 for patients with and without CLEC2A expression, respectively). Given high rates of co-occurring KMT2A-r AML, particularly high-risk fusions (MLLT10 and MLLT4), we evaluated clinical outcomes within the KMT2A-r cohort as these patients have poor EFS and OS. KMT2A-r patients with CLEC2A had significantly worse EFS (24% vs. 39%, p=0.0063) and OS (38% vs. 60%, p=0.0002). Within the KMT2A-r cohort, high rates of relapse were seen for patients with CLEC2A expression compared to those without CLEC2A (72% vs. 55%, p=0.0039). Here we describe CLEC2A, a novel AML-restricted cell surface target that is an ideal immunotherapeutic target. CLEC2A is highly expressed in KMT2A-r AML, entirely absent in normal hematopoietic cells, directly and causally linked to the KMT2A fusion, and can be used for target-directed cytotoxicity.

T-Cells Immune Cluster Analysis Using Cytof Identifies Unique Patient Subgroups with TP53 Mutation and Predicts Complete Remission (CR) in Patients with Acute Myeloid Leukemia (AML) Receiving Venetoclax Plus Hypomethylating Agent Therapy

Introduction The advent of venetoclax (VEN) based chemotherapy has revolutionized the treatment of elderly acute myeloid leukemia (AML), with improvement in remission rates from historical 10%-20% to 60%-70%. However, remission duration is short-lived in patients (pts) with high-risk AML with consequent dismal outcome. As our knowledge of genomic landscape of AML expands and more immunotherapies become available, exploration of the immune tumor microenvironment (TME) of AML may allow identification of subsets of pts that may benefit from immune based therapy. We conducted an investigator initiated prospective study to evaluate TME of pts receiving VEN plus hypomethylating agent (HMA; azacitidine or decitabine) combination. We hypothesize that these pts will cluster into immune-infiltrated vs. immune-depleted sub-types by high-definition immune cell profiling. Methods In 26 newly diagnosed AML pts, who were planned to receive VEN plus HMA therapy, we isolated bone marrow (BM) and peripheral blood (PB) derived mononuclear cells (MC) by density gradient centrifugation and cryopreserved. We performed CyTOF (cytometry time-of-flight) analysis to study their immune signatures. Thawed cells were labeled with metal-conjugated antibodies that bind CD33, CD196, CD19, CD127, CD11b, CD4, IgD, CD11c, CD14, CD56, CD123, TCRgd, CD185/CXCR5, CD45RA, CD27, CD28, CD66b, CD183 (CXCR3), CD161, CD45RO, CD197 (CCR7), CD8a, CD25, CD3, CD20, CD38, HLA-DR, CD194 (CCR4), CD57, CD16 and CD45. We performed a Rphenograph clustering on 25 paired BMMC and PBMC samples (one patient only had PBMC) to create 35 unique clusters for each ( Figure 1). Then we broke down the distribution of cluster by two different groupings: pts with TP53 mutation (m) and those who achieve complete remission. Statistical significance of clusters calculated using t-test. We performed Pearson correlation test on the paired BMMC and PBMC samples to assess correlation between BMMC and PBMC samples. Results Baseline characteristics The median age of the pts was 74 years (range, 53-88), and 17 (65%) pts were male. Ten (37%) pts had secondary AML, and 12 (46%) pts had ELN adverse risk disease (n=6; TP53m). There was no significant difference in median age (p= >0.99), proportion of pts with secondary AML (p= 0.64), acute myelomonocytic leukemia (p= 0.14) white blood cell count (WBC) ≥ 100 (10 9/L) (p= >0.99), and frequency of commonly occurring co-mutations ( ASXL1 [p=0.35]), FLT3 ITD [p=0.54], splicing [p=0.29], IDH1/2 [p=0.28], DNMT3A [p=0.29], NPM1 [p=0.29], TET2 [p=0.35]) in TP53m and TP53 wild-type group. Response 12/20 (60%) evaluable pts achieved complete remission with or without count recovery (CR/CRi). There was no significant difference in median age (p= 0.06), proportion of pts with secondary AML (p= 0.16), therapy related AML (p= 0.60) WBC ≥ 100 (10 9/L) (p= 0.40), and frequency of commonly occurring mutations ( TP53 [p=0.60], ASXL1 [p > 0.99]), FLT3 ITD [p=0.54], splicing [p > 0.99], IDH1/2 [p=0.60], DNMT3A [p=>0.99], NPM1 [p=0.61], TET2 [p >0.99]) in CR/CRi and no-CR/CRi group. CyTOF analysis and Rphenograph clustering on 25 paired BMMC and PBMC samples We found significant clustering with high expression of B-cell activated (cluster 2; p= 0.005), NK cell rich (cluster 1; p= 0.005, cluster 35; p= 0.0009), CD8 T-cells (cluster 31; p= 0.01) and CD4 effector cells (cluster 17; p= 0.005) phenotype among TP53m AML ( Table 1). Among pts who achieved CR/CRi, significant clustering with low expression of B-cell activated (cluster 29; p= 0.01), CD8 cytotoxic (cluster 14; p= 0.01) CD4 activated (cluster 21; p= 0.03) phenotypes observed. Correlation between BMMC and PBMC clustering Among clusters which showed significance for a particular phenotype, we performed Pearson correlation (R 2) and observed strong correlation between BMMC and PBMC samples clustering in cluster 1 (R 2 = 0.99), cluster 2 (R 2 = 0.95), cluster 17 (R 2 = 0.96), cluster 21 (R 2 = 0.99) and cluster 29 (R 2 = 0.93). Conclusion In this single institutional study using CyTOF analysis we have identified novel clustering with immune- infiltrated phenotype among pts with TP53m AML and conversely immune-depleted phenotype among AML pts who responded to VEN plus HMA. Moreover, we found strong correlation in CyTOF analysis conducted from BMMC and PBMC samples. Future studies with larger sample size needed to validate our findings.

Li-Fraumeni Syndrome: The Two Faces of a Coin in Myeloid Malignancies

Background Li-Fraumeni Syndrome (LFS) is a rare, autosomal dominant genetic disorder caused by germline mutations in the TP53 tumor suppressor gene. It is characterized by a predisposition to a wide range of cancers, including hematological malignancies. Hematological malignancies account for approximately 3-4% of all LFS-related cancers of which are acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), Myelodysplastic syndromes (MDS) and lymphomas. Method and Results We retrospectively analyzed our clinical data for patients with hematological malignances from the period of Jan 2016-Jan 2022 at the national center for cancer care and research (NCCCR), the only tertiary hematology center in the country. We report on two cases with Li Fraumeni syndrome with acute leukemia and their variable spectrum of the disease manifestations and prognosis. Case 1: 16 years old male known case of rheumatic heart disease presented with double malignancy; left mandible osteosarcoma and lymphoblastic leukemia/ lymphoma concurrently in 2020. For his Leukemia he received BFM*2009 protocol and after remission he underwent surgical resection and received radiotherapy for the osteosarcoma. Patient has family history of breast cancer in his maternal aunt and his paternal great aunt. Germline Genetic testing from buccal swab revealed a pathogenic variant in TP53 gene confirming Li Fraumeni syndrome which is most likely De novo as it was not detected in both parents and his 3 siblings. In 2022 patient presented with pancytopenia and therapy related AML/MDS with complex Karyotype. He was started on salvage therapy and was planned for allogenic stem cell transplant (SCT) from his HLA matched brother, however, unfortunately, patient didn't achieve complete remission thus could not receive the SCT and he passed away one year later in 2023. Case 2: 60-year-old female known case of hypertension who was diagnosed with right breast cancer in 2011 for which she underwent surgery, chemotherapy, and radiotherapy. A year later, patient was diagnosed with left upper lobe squamous cell lung carcinoma, and underwent left upper lobectomy, chemotherapy and radiotherapy. Patient was in remission for 6 years, until 2018 when she presented with pancytopenia and therapy related AML with complex monosomic Karyotype. Due to patient's age and comorbidities, she was unfit for intensive therapy, thus she received hypomethylation agent (Azacitidine) till disease progression and was referred to genetic counseling. Patient has a family history of unknown cancer in her paternal half-sister at the age of 77 and an unknown cancer in her paternal cousin. Genetic testing reveled a mosaic pathogenic variant in TP53 gene present in 17% of the 1137 next generation sequencing reads. Shortly later, patient passed away without confirmatory fibroblast testing. Conclusion: Genetic testing indications for Li-Fraumeni syndrome present challenges as patients with full mutationin TP53 often but not always present with the full spectrum of the disease, whereas patients with mosaic profilepresent with a later onset of the disease and might not be identified early. In both scenarios, patients can be miss-identified and exposed to radiotherapy thus testing criteria warrants further evaluation. Acute Leukemia in Li-Fraumeni syndrome is associated with poor prognosis and usually presents in the context of therapy related myeloid malignancies . Identification of patients with Li-Fraumeni syndrome early on is crucial to avoid exposures to radiotherapy whenever possible thus reducing the risk of subsequent malignancies and the tragic outcomes of the disease. There are unmet clinical needs in Li-Fraumeni syndrome diagnostic and therapeutic approaches which needs further attention.

Clinical outcomes after CPX-351 in patients with high-risk acute myeloid leukemia: A comparison with a matched cohort from the Spanish PETHEMA registry.

CPX-351 is approved for the treatment of therapy related acute myeloid leukemia (t-AML) and AML with myelodysplastic related changes (MRC-AML). The benefits of this treatment over standard chemotherapy has not been addressed in well matched cohorts of real-life patients. Retrospective analysis of AML patients treated with CPX-351 as per routine practice. A propensity score matching (PSM) was used to compare their main outcomes with those observed in a matched cohort among 765 historical patients receiving intensive chemotherapy (IC), all of them reported to the PETHEMA epidemiologic registry. Median age of 79 patients treated with CPX-351 was 67 years old (interquartile range 62-71), 53 were MRC-AML. The complete remission (CR) rate or CR without recovery (CRi) after 1 or 2 cycles of CPX-351 was 52%, 60-days mortality 18%, measurable residual disease <0.1% in 54% (12 out of 22) of them. Stem cell transplant (SCT) was performed in 27 patients (34%), median OS was 10.3 months, and 3-year relapse incidence was 50%. Using PSM, we obtained two comparable cohorts treated with CPX-351 (n = 52) or IC (n = 99), without significant differences in CR/CRi (60% vs. 54%) and median OS (10.3 months vs. 9.1 months), although more patients were bridged to SCT in the CPX-351 group (35% vs. 12%). The results were confirmed when only 3 + 7 patients were included in the historical cohort. In multivariable analyses, SCT was associated with better OS (HR 0.33 95% CI: 0.18-0.59), p < 0.001. Larger post-authorization studies may provide evidence of the clinical benefits of CPX-351 for AML in the real-life setting.

Outcomes after intensive chemotherapy for secondary and myeloid-related changes acute myeloid leukemia patients aged 60 to 75 years old: a retrospective analysis from the PETHEMA registry.

Treatment options for patients with secondary acute myeloid leukemia (sAML) and AML with myeloid-related changes (AMLMRC) aged 60 to 75 years are scarce and unsuitable. A pivotal trial showed that CPX-351 improved complete remission with/without incomplete recovery (CR/CRi) and overall survival (OS) as compared with standard "3+7" regimens. We retrospectively analyze outcomes of 765 patients with sAML and AML-MRC aged 60 to 75 years treated with intensive chemotherapy, reported to the PETHEMA registry before CPX-351 became available. The CR/CRi rate was 48%, median OS was 7.6 months (95% confidence interval [CI]: 6.7-8.5) and event-free survival (EFS) 2.7 months (95% CI: 2-3.3), without differences between intensive chemotherapy regimens and AML type. Multivariate analyses identified age ≥70 years, Eastern Cooperative Oncology Group performance status ≥1 as independent adverse prognostic factors for CR/CRi and OS, while favorable/intermediate cytogenetic risk and NPM1 were favorable prognostic factors. Patients receiving allogeneic stem cell transplant (HSCT), autologous HSCT, and those who completed more consolidation cycles showed improved OS. This large study suggests that classical intensive chemotherapy could lead to similar CR/CRi rates with slightly shorter median OS than CPX-351.

Molecular genetic and cytofluorimetric prognostic factors in the development of acute myeloid leukemia relapse in children after allogeneic hematopoietic stem cell transplantation

Relapse of acute myeloid leukemia (AML) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) remains one of the main causes of reduced long-term survival. Modern methods for predicting the risk of AML relapse after allo-HSCT take into account the data on the pre-transplant level of minimal residual disease (MRD) determined by flow cytometry and molecular biological studies of recurrent genetic abnormalities, which are currently widespread in clinical practice. Recent studies of the expression of genes characteristic of leukemic stem cells (LSCs) have shown prognostic significance for children with AML in relation to treatment response and the risk of relapse. The study of LSC persistence in order to predict the risk of recurrence after allo-HSCT in children with AML in addition to standard MRD detection methods may be of great importance. The aim of the work was to evaluate the impact of MRD status, both using classic methods and taking into account the genes characteristic of LSC, on the results of allo-HSCT in children with AML. The study was approved by the Independent Ethics Committee and the Scientific Council of the I.P. Pavlov First Saint Petersburg State Medical University of Ministry of Healthcare of the Russian Federation. To assess MRD using standard diagnostic methods, we analyzed the data of 95 children with AML in their 1st–2nd remission (cohort 1). MRD status was negative in 67 (70.6 %) patients; in 28 (29.4 %) children, MRD status was positive according to molecular genetic studies and / or immunophenotyping results. For pre-transplant evaluation of the expression of genes characteristic of LSC, we investigated bone marrow samples of 50 patients (cohort 2) using real-time polymerase chain reaction. The DNMT3B, GPR56, CD34, SOCS2, SPINK2, FAM30A, and ABL genes were studied by real-time polymerase chain reaction, followed by calculation of the pLSC6 value using the formula: DNMT3b × 0.189 + GPR56 × 0.054 + CD34 × 0.0171 + SOCS2 × 0.141 + SPINK2 × 0.109 + FAM30A × 0.0516. At the time of allo-HSCT, 37 (74 %) children with AML were in their 1st or 2nd remission of the disease, 13 (26 %) were out of the 1st–2nd remission. With a median follow-up of 5 years in the group of patients with a positive MRD status, determined by standard methods (cohort 1), overall survival (OS) was 67.9 % vs 73.1 % for patients with a negative MRD status (p = 0.83). The cumulative incidence of relapse was 50 % and 22 %, respectively; p = 0.012. When assessing the level of expression of genes characteristic of LSC (cohort 2), a pLSC6 level was above the median in 18/37 (49 %) patients. The results of linear regression showed that the pre-transplant level of expression of genes characteristic of LSC was not associated with the number of blasts/MRD (odds ratio 1.002; 95 % confidence interval 0.979–1.025). One-year OS rates did not differ significantly in children in the 1st–2nd remission of AML, depending on pLSC6 level: 84.2% in patients with low pLSC6 and 72.2 % – with high pLSC6 (p = 0.4), event-free survival in the corresponding groups – 68.4 % and 61.1 %, respectively (p = 0.34). The cumulative incidence of early relapse after allo-HSCT in the group of AML patients with a high pLSC6 score was significantly higher than in children with a low pLSC6 score before allo-HSCT (22 % and 0 %, respectively; p = 0.03). MRD does not have a statistically significant effect on OS. However, MRD positivity before allo-HSCT increases cumulative incidence of relapse. The level of expression of genes characteristic of LSC, determined before allo-HSCT, showed a prognostic significance in relation to the development of early AML relapse after allo-HSCT.

The prognostic signature based on glycolysis-immune related genes for acute myeloid leukemia patients

Acute myeloid leukemia (AML) is widely considered an immunoresponsive malignancy. However, potential association between glycolysis-immune related genes and AML patients’ prognosis has been seldom studied. AML-related data was downloaded from TCGA and GEO databases. We grouped patients according to Glycolysis status, Immune Score and combination analysis, basing on which overlapped differentially expressed genes (DEGs) were identified. The Risk Score model was then established. The results showed that totally 142 overlapped genes were probably correlated with glycolysis-immunity in AML patients, among which 6 optimal genes were screened to construct Risk Score. High Risk Score was an independent poor prognostic factor for AML. In conclusion, we established a relatively reliable prognostic signature of AML based on glycolysis-immunity related genes, including METTL7B, HTR7, ITGAX, TNNI2, SIX3 and PURG.

Evaluation of 5-Day Vs. 10-Day Decitabine in Combination with Venetoclax in Newly Diagnosed AML Patients Ineligible for Induction Chemotherapy

BACKGROUND. Standard dosing of decitabine 20mg/m 2 daily given over 5 days in combination with the Bcl-2 inhibitor venetoclax (5-day DEC/VEN), in elderly or chemotherapy ineligible patients with Acute Myeloid Leukemia (AML), induces both a high rate of complete remission (CR/CRi rate 71%) and prolonged survival (median OS 14.2 months). (DiNardo CD et al. Blood. 2019) Extending the duration of decitabine treatment to 10-days (10-day DEC/VEN) may increase efficacy of the combination. In a phase 2 study of newly diagnosed chemotherapy ineligible AML patients aged ≥ 60 years, 10-day DEC/VEN led to a high CR/CRi rate of 89%, with 67% of these being negative for measurable residual disease and a median OS of 18 months. (DiNardo, CD et al. Leukemia. 2020). As a prospective clinical trial comparing efficacy of the different dosing schedule of decitabine in combination with venetoclax is unlikely to be performed, we conducted a single center retrospective analysis comparing 5-day and 10-day DEC/VEN outcomes. METHODS. We retrospectively studied patients with newly diagnosed AML receiving either 5-day or 10-day DEC/VEN between January 2016 to May 2022 at City of Hope National Medical Center. Patients receiving prior AML therapies including hypomethylating agents were excluded. Responses were evaluated per the ELN criteria (Dohner, H et al. Blood. 2017) Measurable residual disease (MRD) flow cytometry was performed at the University of Washington. Patient characteristics were summarized by frequency and associations between overall response and patient and disease characteristics were tested by Fisher's exact test. Overall Survival (OS) was evaluated by the Kaplan-Meier method and the difference between groups was determined by log-rank test. All statistical analyses were performed using R software. RESULTS. 139 patients were included: 48% (n=67) treated with 5-day decitabine and 52% (n=72) treated with 10-day decitabine. The baseline characteristics of the patients are summarized in table 1. At baseline, 5-day decitabine + venetoclax treated patients were older with a median of age of 73 vs. 70 years (p=0.01). The 5-day and 10-day groups were otherwise well balanced with no significant difference in AML type, ELN risk or adverse molecular features. Rates of CR/CRi, CR/CRi, MRD negativity, and the percentage of responders proceeding to allogeneic stem cell transplant were similar between regimens (Table 1). There was also no difference in 8-week mortality (p = 0.44). With a median follow-up of 12 months for responding patients, the median OS (figure 1) and RFS for 5-day and 10-day DEC/VEN were 14 vs. 27 months, p=0.091 and 12.9 vs. 14.3 months, p= 0.75 respectively. In subgroup analysis, patients without therapy- related AML (non-tAML) receiving the 10-day-regimen had a longer median OS (42.5 vs. 14.6 months, p= 0.03). No other subgroups, including those with high-risk molecular (eg.TP53) or cytogenetic risk factors (e.g. complex karyotype) demonstrated a significant difference between the 5 and 10-day DEC/VEN regimen. CONCLUSION. Among newly diagnosed AML patients receiving venetoclax in combination with either 5-day or 10-day decitabine, there was no significant difference in response (CR/CRi or MRD negativity) or survival outcomes (OS or RFS). Although a significant difference was not observed, 10-day DEC/VEN led to an increased median OS, which may reach significance with more patients and longer follow-up. Additionally, the 10-day DEC/VEN regimen may benefit certain subpopulations; it demonstrated longer median OS in patients without t-AML. Further analysis to confirm a survival benefit among patients without t-AML receiving either 5-day or 10-day DEC/ VEN is needed and will be presented at ASH. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Outcomes of Allogeneic Haematopoietic Stem Cell Transplantation for Therapy-Related Myeloid Neoplasms Following Treatment for Myeloma

Introduction As the outcomes for patients with multiple myeloma continue to improve, therapy-related myeloid neoplasms (t-MN) are increasingly recognised as a treatment complication. The prognosis of tMDS is poor with life expectancy typically less than a year1. There is limited data on the management of t-MN secondary to myeloma therapies. We performed a registry study on the outcomes of patients who underwent an allogeneic haematopoietic stem cell transplant (allo-HCT) for both therapy related myelodysplastic syndrome (t-MDS) and acute myeloid leukemia (t-AML) following treatment for myeloma. Methods The European Society for Blood and Marrow Transplantation (EBMT) registry was used to identify 157 patients with t-MN across 96 centres between 2006-2018. The median age of patients was 62.1 years (57.6-66.7), with 63.7% male and 36.5% female. 100 patients were transplanted for t-AML and 57 for t-MDS with a median time from myeloma diagnosis to time of allo-HCT of 72.6 (46.1-102.9) months. The median time from t-MN diagnosis to allo-HCT was 6.4 (3.9-9.4) months. We examined overall survival (OS), progression free survival (PFS) relapse and non-relapse mortality (NRM) and myeloma recurrence. The median follow-up time was 64.9 months. Results The median survival time for all patients was 14.3 months. OS at 12- and 60 months was 55% and 27% respectively and PFS at 12- and 60 months was 45% and 24% respectively. The incidence of relapse of tMN at 12- and 60 months was 35% and 45% respectively, with NRM 20% and 31% respectively. The commonest cause of death was infection (36.3% n=37) followed by disease relapse/progression (34.3% n=35). With respect to t-MN disease status 59% of patients with t-MN were in a complete remission at time of transplant. Cytogenetic results were available for all patients with 67.6% abnormal, 26.7% normal and failed/not done in 5.7% cases. IPPS-R was available in 58% of cases; Good 17(29.3%), intermediate 13(22.4%), poor 13(22.4%) and very poor 15(25.9%). Data on status of myeloma at time of allo-HCT was available in only 50 patients (CR 38%, VGPR 22%, PR 10%, stable disease 11% and progressive disease 8%). The majority of patients underwent matched unrelated transplant 79%(124) versus sibling allo 21% (n33). The majority had reduced intensity conditioning (RIC 70.3%, standard 29.7%, missing 1.3%). 130 patients had undergone prior autologous transplant (auto) for myeloma with 55.4% having one auto, 21% two autos, 5.1% three autos and 1.3% four autos. With respect to the number of lines of myeloma therapy prior to transplant; 1 line (29.4%), 2 (30.9%), 3 (16.9%), 4 (8.8%) and >4 (14%). 120 patients received chemotherapy for myeloma prior to their t-MN diagnosis, however data was missing in the remainder of patients in this study. Data on myeloma recurrence was available in 31.2% (49 patients), at 12- and 60 months myeloma recurrence was 4% and 12% respectively with myeloma NRM at 12- and 60 months 33% and 57%. Univariate analysis failed to show any statistically significant difference in outcomes with respect to t-MDS vs t-AML, t-MN disease status, Karnofsky performance status, conditioning or donor source. Multivariate analysis was not possible due to the small numbers within this study. Further analysis of this data will be undertaken. Conclusion This study indicates that allo-HCT for t-MN following treatment for myeloma is a viable treatment option. The myeloma recurrence in this study was low and further strategies to reduce t-MN relapse and NRM are needed. References; Bhatia R, Deeg HJ. Treatment-Related Myelodysplastic Syndrome - Molecular Characteristics and Therapy. Curr Opin Hematol.2011;18(2):77-82.

Donor Derived Myelodysplastic Syndrome after Sex Matched Haploidentical Allograft for AML with Monosomy 7 in Both Diseases: Importance of Donor Chimerism Assessment By Two Independent Techniques

Donor-derived leukaemia is a rare outcome in patients following allogeneic hematopoietic cell transplant (HCT) accounting for ~ 0.1-5% of relapsed cases although the true incidence is likely underestimated. A 39-year-old male patient with a history of ulcerative colitis treated with mercaptopurine was subsequently diagnosed with therapy related acute myeloid leukaemia (AML) and karyotype showed 45,XY,-7. The patient achieved remission after two cycles of treatment and proceeded to a paternal haploidentical HCT. Complete remission with no detectable disease by flow cytometry and fluorescent in-situ hybridisation (FISH) was achieved by day 40 of transplantation with whole blood and T-cell chimerism of 97% as assessed by routine short tandem repeat (STR) chimerism analysis. With withdrawal of immunosuppression this increased to 100% in both fractions. At 11 months post-transplant evidence of low-level cytogenetic relapse emerged with monosomy 7 seen by FISH in 15/200 interphase cells. Chimerism remained stable at 99% with aspirate findings indicating 2-3% blasts and some trilineage dysplasia. Two subsequent marrows showed persistence of monosomy 7 by FISH with full donor STR chimerism. Although a possibility of donor derived myelodysplastic syndrome (DD-MDS) was considered, due to the monosomy 7, donor lymphocyte infusion (DLI) therapy was administered at 16 and 19 months post-transplant. Monosomy 7 persisted on FISH and standard cytogenetic analysis. To confirm validity of STR chimerism and to distinguish between DD-MDS and relapse of original disease, HLA typing by next generation sequencing (NGS) to exclude HLA loss related relapse and potentially guide rescue decisions was performed alongside NGS chimerism assessment. NGS chimerism utilises genetic markers with population independent discriminative power distributed across 17 chromosomes and further selected to allow sensitive detection combined with accurate and precise quantification of mixed chimerism. NGS chimerism is demonstrated to be sensitive enough to detect a minor DNA contributor comprising down to 0.5% of the sample, detection of low level mixed chimerism and therefore clinical utility in better predicting relapse compared to conventional techniques. Original stored donor cells were tested and showed no evidence of monosomy 7 by FISH. After confirming DD-MDS a second HCT was considered. Originally unsuccessful unrelated donor search was re-opened and wide family donor search initiated. With azacytidine therapy the patient achieved a short lived cytogenetic remission, but subsequently relapsed and progressed to AML with karyotype of 45,XY,-7,der(16)t(11;16)(q13;p13.1)[9]/46,XY[1] and died 33 months after HCT prior to initiation of a second HCT. This case was complicated by the presence of monosomy 7 detected by karyotyping in both the initial diagnostic t-AML and in the DD MDS. Monosomy 7 is the most common abnormality found in DD-MDS but a review of the literature available does not seem to indicate any cases where it was also present in the patient's initial diagnostic sample as a separate disease entity. The pathogenesis of DD-MDS remains unclear. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

The Impact of Pre-Transplant Extramedullary Disease on the Outcome of Allogeneic Hematopoietic Cell Transplantation for Acute Myeloid Leukemia in Children- on Behalf of PDWP/EBMT

Background Extramedullary disease (EMD), including central nervous system (CNS) and myeloid sarcoma, is a rare presentation in Acute Myeloid Leukemia (AML). We studied the effect of EMD (CNS involvement only and other EMD ± CNS) compared to isolated BM involvement in the outcomes following hematopoietic stem cell transplantation for AML in children. Methods An EBMT registry-based retrospective study to assess the impact of extramedullary disease (EMD) in children (de novo AML, age <18y at transplant, period 2008-2016) with AML who underwent a first allogeneic hematopoietic cell transplant (allo-HCT) with non-TBI conditioning regimen. Outcomes of interest included: leukemia-free survival (LFS), overall survival (OS), relapse incidence (RI) and non-relapse mortality (NRM). Patients were grouped into three categories: BM involvement only (Group A), BM+CNS involvement only (Group B) and BM+ other EMD ± CNS (Group C). Patients with Down syndrome related AML and t(15:17) AML were excluded. Results A total of 958 (529 males) patients met study criteria, including 685 (71.5%) Group A, 135 (14.1%) Group B and 138 (14.4%) Group C patients. Group C patients were transplanted at a younger age (median age at transplant 4.8y vs 9.2y and 6.6y for Group A, B respectively, p <0.001). A higher proportion of Group A patients were in CR1 (70.8%) compared to Group B (63.7%) and C (60.1%) patients. The three groups were comparable regarding the distribution of high and standard risk cytogenetics. Fifty seven percent of the patients received an unrelated donor and 35% a matched sibling/family donor. The median post HCT follow-up was 5.4 years (IQR: 5.2-5.7y). Five years LFS, OS, RI and NRM were 62%, 68.4%, 26.4% and 11.6%, respectively in the entire cohort. Multivariate analysis showed higher RI in Group C compared to Group A (HR= 1.45 (1.01-2.06) p=0.04) and a non-significantly different NRM (HR: 0.61 (0.29-1.29), p=0.2). There was no difference in RI in Group B compared to Group A (Hazard Ratio (HR) : 1.12 (0.76-1.64). In the multivariate analysis patients with EMD (CNS only or EMD+/- CNS) had no significant difference in LFS, OS and NRM. The Hazard ratio of LFS, OS and NRM for Group B and C compared to Group A was: HR=1.13, p= 0.46; HR=0.96, p=0.82; HR=1.09, p=0.76 and HR=1.19, p=0.28; HR=0.96, p = 0.82; HR=0.61, p=0.2, respectively. Further 116 patients underwent hematopoietic stem cell transplantation with active disease. Eighty-nine patients had BM involvement only (group A), 12 patients had BM and CNS disease (Group B) and 15 patients had BM and other sites of EMD +/- CNS (Group C). In this cohort, the 4y-OS was 37% in Group A, 25% in Group B of and 38% in Group C. Four-year LFS was 31%, 25% and 39% in Group A, Groups B and Group C respectively. Conclusions Our findings suggest that children with AML and BM with EMD ± CNS involvement (group C) have a higher incidence of relapse after HCT compared to those with BM only or BM+CNS only disease. However, the presence of EMD with CNS involvement or other extramedullary disease did not have a statistically significant impact on overall survival, leukemia-free survival or non-relapse mortality. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Outcomes with Venetoclax Dose De-Escalation and Variable Cycle Intervals: A Real-World AML Survival Analysis

The treatment paradigm for acute myeloid leukemia (AML) includes regimens each specified for a target population. Unfit patients (pts) and those of older age are often limited in their treatments considering the toxicity profile associated with traditional induction regimens. Venetoclax (Ven), a potent BCL-2 inhibitor, is frequently administered for older adults who are deemed ineligible for more intensive chemotherapies. However, its clinical use remains a challenge due to its severe myelosuppressive effects. Despite the generally accepted clinical practice of delaying the next cycle or reducing the total Ven dosage per cycle, there is minimal data evaluating the effectiveness and resource needs under these circumstances. The objective of our analysis was to review the clinical course and identify survival rates in our pts undergoing treatment with Ven for AML. We retrospectively reviewed the clinical and molecular data of newly diagnosed and relapsing/refractory AML pts admitted to North Shore University Hospital for induction treatment with a hypomethylating agent (HMA) plus Ven from Nov 2019 to May 2022. Pts were categorized into subgroups based on their median number of days on Ven per cycle excluding induction cycle (≤ 14 days, 21 days, 28 days) as well as the median number of interval days between cycles (< 35 days or ≥ 35 days). Pts who did not complete 2 cycles were excluded from subgroup analyses. Median progression-free survival (mPFS) was defined as the time from Ven initiation to disease progression or death from any cause. PFS was analyzed with the Kaplan-Meier (KM) method and log rank testing. One-way analysis of variance and t-test was used to compare clinical interventions in each subgroup. A total of 53 pts underwent induction therapy with Ven + HMA: 79.2% frontline and 20.1% relapsed/refractory. The median age was 76 years-old (range 60-91). Of these 53 pts, 62.2% (n=33) were de novo, 24.5% (n=13) had AML with myelodysplastic related changes and 13.2% had therapy related AML (n=7). Utilizing the 2022 ELN risk categories (Dohner et al, Blood 2022), < 1% (n=3) were favorable risk while 33.9% (n=18) were intermediate risk and 60.3% (n=32) were adverse risk. All prognostic indicators were evenly dispersed among the subgroups. 33.9% of pts (n=18) died within their induction treatment admission. The mPFS was not reached in the subgroup of pts receiving ≤ 14 days of Ven/cycle while it was 10.8 months in pts receiving 21 days of Ven/cycle and 11.9 months for pts receiving 28 days of Ven/cycle. Although there was no significant difference between KM estimates when comparing number of days on Ven/cycle, there was a trend favoring ≤14 days of Ven/cycle. Patients who had ≥ 35 days interval between cycles had a mPFS of 11.9 months versus 8.5 months in pts who had < 35 days intervals. However, their KM estimates were not significantly different (p > 0.05). Pts included in subgroup analysis, received an average of 6.7 cycles and required an average of 2.6 hospital admissions while on a Ven regimen. Pts received an average of 4.8 pRBC units and 6.1 platelet units per cycle. Although there was a trend favoring reduction in Ven dosing (≤ 14 day subgroup), there was no significant difference in number of cycles, number of hospital admissions and transfusion requirements between each subgroup analysis (p > 0.05). This retrospective analysis demonstrates no difference in Ven outcomes with dose de-escalation and increased interval time between cycles in pts requiring such adjustments. The mPFS of each subgroup analyses was also similar compared to the mPFS (14.7 months) reported in the phase 3 clinical trial leading to its FDA approval (DiNardo et al, NEJM 2020; NCT02993523). Interestingly, the subgroup with a median dosing of ≤ 14 days of Ven/cycle had yet to reach a mPFS at 18 months. Also, the review of our patients’ clinical courses progress demonstrated a trend in decreased transfusion requirements and hospitalizations with lower Ven dosing although not statistically significant. Our analysis suggests that further larger retrospective and prospective enquiries are needed to evaluate the potential benefit of Ven dose de-escalation not only in terms of PFS and treatment related complications, but also for quality of life. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Prognostic Value of FLT3-Internal Tandem Duplication Residual Disease in Acute Myeloid Leukemia.

The applicability of FLT3-internal tandem duplications (FLT3-ITD) for assessing measurable residual disease (MRD) in acute myeloid leukemia (AML) in complete remission (CR) has been hampered by patient-specific duplications and potential instability of FLT3-ITD during relapse. Here, we comprehensively investigated the impact of next-generation sequencing (NGS)-based FLT3-ITD MRD detection on treatment outcome in a cohort of patients with newly diagnosed AML in relation to established prognostic factors at diagnosis and other MRD measurements, ie, mutant NPM1 and multiparameter flow cytometry. In 161 patients with de novo FLT3-ITD AML, NGS was performed at diagnosis and in CR after intensive remission induction treatment. FLT3-ITD MRD status was correlated with the cumulative incidence of relapse and overall survival (OS). NGS-based FLT3-ITD MRD was present in 47 of 161 (29%) patients with AML. Presence of FLT3-ITD MRD was associated with increased risk of relapse (4-year cumulative incidence of relapse, 75% FLT3-ITD MRD v 33% no FLT3-ITD MRD; P < .001) and inferior OS (4-year OS, 31% FLT3-ITD MRD v 57% no FLT3-ITD MRD; P < .001). In multivariate analysis, detection of FLT3-ITD MRD in CR confers independent prognostic significance for relapse (hazard ratio, 3.55; P < .001) and OS (hazard ratio 2.51; P = .002). Strikingly, FLT3-ITD MRD exceeds the prognostic value of most generally accepted clinical and molecular prognostic factors, including the FLT3-ITD allelic ratio at diagnosis and MRD assessment by NGS-based mutant NPM1 detection or multiparameter flow cytometry. NGS-based detection of FLT3-ITD MRD in CR identifies patients with AML with profound risk of relapse and death that outcompetes the significance of most established prognostic factors at diagnosis and during therapy, and furnishes support for FLT3-ITD as a clinically relevant biomarker for dynamic disease risk assessment in AML.

Recommendations for laboratory testing of UK patients with acute myeloid leukaemia.

Recommendations for laboratory testing of UK patients with acute myeloid leukaemia.