Pathologic, cytogenetic, and molecular features of acute myeloid leukemia with megakaryocytic differentiation: A report from the Children's Oncology Group.

2023

e30251

02/2023

1545-5017

BACKGROUND: Acute myeloid leukemia (AML) with megakaryocytic differentiation (AMkL) is a rare subtype of AML more common in children. Recent literature has identified multiple fusions associated with this type of leukemia.

METHODS: Morphology, cytogenetics, and genomic sequencing were assessed in patients from Children's Oncology Group trials AAML0531 and AAML1031 with central-pathology review confirmed non-Down syndrome AMkL. The 5-year event-free survival (EFS), overall survival (OS), and RR were evaluated in these AMkL subcategories.

RESULTS: A total of 107 cases of AMkL (5.5%) were included. Distinct fusions were identified in the majority: RBM15::MRTFA (20%), CBFA2T3::GLIS2 (16%), NUP98 (10%), KMT2A (7%), TEC::MLLT10 (2%), MECOM (1%), and FUS::ERG (1%); many of the remaining cases were classified as AMkL with (other) myelodysplasia-related changes (MRC). Very few cases had AML-associated somatic mutations. Cases with CBFA2T3::GLIS2 were enriched in trisomy 3 (p = .015) and the RAM phenotype, with associated high CD56 expression (p < .001). Cases with NUP98 fusions were enriched in trisomy 6 (p < .001), monosomy 13/del(13q) (p < .001), trisomy 21 (p = .026), and/or complex karyotypes (p = .026). While different 5-year EFS and OS were observed in AMkL in each trial, in general, those with CBFA2T3::GLIS2 or KMT2A rearrangements had worse outcomes compared to other AMkL, while those with RBM15::MRTFA or classified as AMkl-MRC fared better. AMkL with NUP98 fusions also had poor outcomes in the AAML1031 trial.

CONCLUSION: Given the differences in outcomes, AMkL classification by fusions, cytogenetics, and morphology may be warranted to help in risk stratification and therapeutic options.

10.1002/pbc.30251

Pediatr Blood Cancer

36789545

SAMHD1 Single Nucleotide Polymorphisms Impact Outcome in Children with Newly Diagnosed Acute Myeloid Leukemia.

2023

01/2023

2473-9537

Cytarabine arabinoside (Ara-C) has been the cornerstone of AML chemotherapy for decades. Following cellular uptake, it is phosphorylated into its active triphosphate form (Ara-CTP), which primarily exerts its cytotoxic effects by inhibiting DNA synthesis in proliferating cells. Interpatient variation in the enzymes involved in the Ara-C metabolic pathway have been shown to impact intracellular abundance of Ara-CTP and thus its therapeutic benefit. Recently, SAMHD1 (SAM and HD domain containing deoxynucleoside triphosphate triphosphohydrolase 1) has emerged to play a role in Ara-CTP inactivation, development of drug resistance and consequently, clinical response in AML. Despite this the impact of genetic variations in SAMHD1 on outcome in AML has not been investigated in depth. In this study, we evaluated 25 single nucleotide polymorphisms (SNPs) within SAMHD1 gene for association with clinical outcome in 400 newly diagnosed pediatric AML patients from two clinical trials- AML02 and AML08. Three SNPs, rs1291128, rs1291141, and rs7265241 located in the 3' region of SAMHD1 were significantly associated with at least one clinical outcome endpoint: minimal residual disease (MRD) after induction I, event free survival (EFS), or overall survival (OS) in the two cohorts. In an independent cohort of patients from COG-AAML1031 trial (n=854), rs7265241 A>G remained significantly associated with EFS and OS. In multivariable analysis, adjusting for other prognostic factors such as race, age, risk group, and white blood cell count, all the SNPs remained independent predictors of clinical outcome endpoints. These results highlight the relevance of the SAMHD1 pharmacogenomics in context of response to Ara-C in AML and warrants the need for further validation in expanded patient cohorts.

10.1182/bloodadvances.2022009088

Blood Adv

36689724

Clinical relevance of proteomic profiling in de novo pediatric acute myeloid leukemia: a Children's Oncology Group study.

2022

2022 Jan 13

1592-8721

<p>Pediatric acute myeloid leukemia (AML) remains a fatal disease for at least 30% of patients, stressing the need for improved therapies and better risk stratification. As proteins are the unifying feature of (epi)genetic and environmental alterations, and are often targeted by novel chemotherapeutic agents, we studied the proteomic landscape of pediatric AML. Protein expression and activation levels were measured in 500 bulk leukemic patient samples and 30 control CD34+ samples, using the reverse phase protein arrays with 296 strictly validated antibodies. The multi-step "MetaGalaxy" analysis methodology was applied and identified nine protein expression signatures (PrSIG), based on strong recurrent protein expression patterns. PrSIGs were associated with cytogenetics and mutational state, and with both favorable or unfavorable prognosis. Analysis based on treatment (i.e., ADE vs. ADE plus bortezomib (ADEB)) identified three PrSIGs that did better with ADEB vs. ADE. When PrSIGs were studied in the context of genetic subgroups, PrSIGs were independently prognostic after multivariate analysis, suggesting a potential value for proteomics in combination with current classification systems. Proteins with universally increased (n=7) or decreased (n=17) expression were observed across PrSIGs. Expression of certain proteins significantly differentially expressed from normal could be identified, forming a hypothetical platform for personalized medicine.</p>

10.3324/haematol.2021.279672

Haematologica

35021602

CBFB-MYH11 Fusion Transcripts Distinguish Acute Myeloid Leukemias with Distinct Molecular Landscapes and Outcomes.

2021

2021 Sep 21

2473-9537

<p>Patients with inv(16)/CBFB-MYH11 AML are considered favorable risk, however, nearly one-third relapse despite intensive therapy. Despite efforts to define risk groups within this favorable risk cohort, CBFB-MYH11 AML patients continue to be treated as a uniform cohort. Through transcriptome sequencing of 186 patients with inv(16) AML, we demonstrate that fusion junction breakpoints (exon 5-exon 33 versus other) are highly associated with outcome. The presence of exon 17 KIT mutations provides additional prognostic significance. Additionally, we provide insights into the transcriptional landscapes that differentiate these distinct CBFB-MYH11 AML subtypes. Children's Oncology Group trials include CCG-2961 (registered at www.clinicaltrials.gov as NCT00002798), AAML03P1 (NCT00070174), AAML0531 (NCT00372593), and AAML1031 (NCT01371981).</p>

10.1182/bloodadvances.2021004965

Blood Adv

34547772

CEBPA bZip Mutations are Associated with Favorable Prognosis in de novo AML: A Report from the Children's Oncology Group.

2021

2021 May 05

1528-0020

<p>Bi-allelic CEBPA mutations are associated with favorable outcomes in AML. We evaluated the clinical and biologic implications of CEBPA-bZip mutations in childhood/young adult newly diagnosed AML. CEBPA-bZip mutation status was determined in 2,958 AML patients enrolled on COG trials (NCT00003790, NCT0007174, NCT00372593, NCT01379181). Next generation sequencing (NGS) was performed in 1,863 patients, 107 with CEBPA mutations, to characterize the co-occurring mutations. CEBPA mutational status was correlated with disease characteristics and clinical outcomes. CEBPA-bZip mutations were identified in 160/2958 (5.4%) patients, with 132 (82.5%) harboring a second CEBPA mutation (CEBPA-dm) and 28 (17.5%) with a single CEBPA-bZip only. The clinical and laboratory features of the two CEBPA cohorts were very similar. CEBPA-dm and CEBPA-bZip patients experienced identical event-free survival (EFS) of 64% and similar overall survival (OS) of 81% and 89%, respectively (p=0.259); this compared favorably to EFS and OS in CEBPA wild type (CEBPA-WT) of 46% and 61%, respectively (both p&lt;0.001). Transcriptome analysis demonstrated similar expression profiles for CEBPA-bZip and CEBPA-dm cases. Comprehensive NGS of CEBPA-mutant cases identified co-occurring CSF3R and GATA2 mutations in 13.1% and 21.5% of patients, respectively. Patients with dual CEBPA/CSF3R mutations had an EFS of 17% vs. 63% for CEBPA-mutant/CSF3R-WT (p&lt;0.001) with a corresponding relapse rate (RR) of 83% vs. 22%, respectively (p&lt;0.001); GATA2 co-occurrence did not impact outcome. CEBPA bZip domain mutations are associated with favorable clinical outcomes, regardless of mono or bi-allelic status. Co-occurring CSF3R and CEBPA mutations are associated with a high RR and nullifies the favorable prognostic impact of CEBPA mutations.</p>

10.1182/blood.2020009652

Blood

33951732

Acute erythroid leukemia is enriched in NUP98 fusions: a report from the Children's Oncology Group.

2020

6000-6008

2020 Dec 08

2473-9537

<p>Acute erythroid leukemia (AEL) is a rare subtype of acute myeloid leukemia (AML) primarily affecting older adults and was previously classified into erythroid/myeloid and pure erythroid subtypes. In this pediatric AEL study, we evaluated morphologic, immunophenotypic, cytogenetic, molecular, and clinical data of 24 (1.2%) cases from all cases undergoing central pathology review in Children's Oncology Group trials AAML0531 and AAML1031. Of 24 cases, 5 had a pure erythroid phenotype, and 19 had an erythroid/myeloid phenotype. NUP98 fusions were highly enriched in patients with AEL, occurring in 7 of 22 cases for which molecular data were available (31.8% vs 6.7% in other AML subtypes). Of 5 cases of pure erythroid leukemias (PELs), 3 had NUP98 fusions, and 4 had complex karyotypes. Erythroid/myeloid leukemias were reclassified by using the 2017 World Health Organization hematopathology classification as: myelodysplastic syndrome (MDS) with excess blasts-1 (n = 3), MDS with excess blasts-2 (n = 7), AML (nonerythroid, n = 5), and unknown MDS/AML (n = 4); the 5 cases of nonerythroid AML included 1 with an NUP98-NSD1 fusion, 2 with myelodysplasia-related changes, and 1 with a complex karyotype. Three cases of MDS with excess blasts-2 also had NUP98 rearrangements. WT1 mutations were present in 5 of 14 cases, all erythroid/myeloid leukemia. Outcomes assessment revealed statistically poorer overall survival (5-year, 20% ± 36% vs 66% ± 23%; P = .004) and event-free survival (5-year, 20% ± 36% vs 46% ± 23%; P = .019) for those with PEL than those with erythroid/myeloid leukemia. Our study supports that AEL is a morphologically and genetically heterogeneous entity that is enriched in NUP98 fusions, with the pure erythroid subtype associated with particularly adverse outcomes.</p>

10.1182/bloodadvances.2020002712

Blood Adv

33284945

Comprehensive Transcriptome Profiling of Cryptic CBFA2T3-GLIS2 Fusion-positive AML Defines Novel Therapeutic Options - A COG and TARGET Pediatric AML Study.

2019

2019 Nov 12

1078-0432

<p><strong>PURPOSE: </strong>A cryptic inv(16)(p13.3q24.3) encoding the CBFA2T3-GLIS2 fusion is associated with poor outcome in infants with acute megakaryocytic leukemia. We aimed to broaden our understanding of the pathogenesis of this fusion through transcriptome profiling.</p>

<p><strong>EXPERIMENTAL DESIGN: </strong>Available RNA from children and young adults with de novo AML (N=1,049) underwent transcriptome sequencing (mRNA and miRNA). Transcriptome profiles for those with the CBFA2T3-GLIS2 fusion (N=24) and without (N=1,025) were contrasted to define fusion-specific miRNAs, genes, and pathways. Clinical annotations defined distinct fusion-associated disease characteristics and outcomes.</p>

<p><strong>RESULTS: </strong>The CBFA2T3-GLIS2 fusion was restricted to infants &lt; 3 years-old (p&lt;0.001) and presence of this fusion was highly associated with adverse outcome (p&lt;0.001) across all morphological classifications. Further, there was a striking paucity of recurrent cooperating mutations and transduction of cord blood stem cells with this fusion was sufficient for malignant transformation. CBFA2T3-GLIS2 positive cases displayed marked up-regulation of genes with cell membrane/extracellular matrix localization potential, including NCAM1 and GABRE. Additionally, miRNA profiling revealed significant over-expression of mature miR-224 and miR-452, which are intronic miRNAs transcribed from the GABRE locus. Gene-set enrichment identified dysregulated Hippo, TGFβ, and hedgehog signaling, as well as NCAM1 (CD56) Interaction pathways. Therapeutic targeting of fusion-positive leukemic cells with CD56-directed ADC caused significant cytotoxicity in leukemic blasts.</p>

<p><strong>CONCLUSIONS: </strong>The CBFA2T3-GLIS2 fusion defines a highly refractory entity limited to infants that appears to be sufficient for malignant transformation. Transcriptome profiling elucidated several highly targetable genes and pathways, including the identification of CD56, providing a highly plausible target for therapeutic intervention.</p>

10.1158/1078-0432.CCR-19-1800

Clin. Cancer Res.

31719049

Functional Properties of KIT Mutations are Associated with Differential Clinical Outcomes and Response to Targeted Therapeutics in CBF Acute Myeloid Leukemia.

2019

2019 Jun 10

1078-0432

<p><strong>PURPOSE: </strong> mutations (+) are common in core binding factor (CBF) AML and have been associated with varying prognostic significance. We sought to define the functional and clinical significance of distinct mutations in CBF pediatric AML.</p>

<p><strong>EXPERIMENTAL DESIGN: </strong>Following transfection of exon 17 (E17) and exon 8 (E8) mutations into HEK293 and Ba/F3 cells, KIT phosphorylation, cytokine independent growth, and response to tyrosine kinase inhibitors (TKI) were evaluated. Clinical outcomes of patients treated on COG AAML0531 (NCT01407757), a phase III study of gemtuzumab ozogamicin (GO), were analyzed according to mutation status (+ vs wild type (-)) and mutation location (E8 vs. E17).</p>

<p><strong>RESULTS: </strong> mutations were detected in 63/205(31%) patients; 22 (35%) involved only E8, 32(51%) only E17, 6(10%) both exons, and 3(5%) alternative exons. Functional studies demonstrated that E17, but not E8, mutations result in aberrant KIT phosphorylation and growth. TKI exposure significantly impacted growth of E17, but not E8, transfected cells. + CBF AML patients had comparable overall survival (OS) to that of - (78%, vs. 81%, p=0.905) but higher relapse rates (RR 43% vs. 21%, p=0.005). E17 + outcomes were inferior to patients [disease free survival (DFS) 51% vs. 73%, p=0.027; RR 21% vs. 46%, p=0.007)] although GO abrogated this negative prognostic impact. E8 mutations lacked significant prognostic impact and GO failed to significantly improve outcome.</p>

<p><strong>CONCLUSIONS: </strong>E17 mutations impact prognosis in CBF AML, as well as response to GO and TKIs, thus clinical trials utilizing both agents should be considered for + patients.</p>

10.1158/1078-0432.CCR-18-1897

Clin. Cancer Res.

31182436

MicroRNA Expression-Based Model Indicates Event-Free Survival in Pediatric Acute Myeloid Leukemia.

2017

JCO2017747451

2017 Oct 25

1527-7755

<p>Purpose Children with acute myeloid leukemia (AML) whose disease is refractory to standard induction chemotherapy therapy or who experience relapse after initial response have dismal outcomes. We sought to comprehensively profile pediatric AML microRNA (miRNA) samples to identify dysregulated genes and assess the utility of miRNAs for improved outcome prediction. Patients and Methods To identify miRNA biomarkers that are associated with treatment failure, we performed a comprehensive sequence-based characterization of the pediatric AML miRNA landscape. miRNA sequencing was performed on 1,362 samples-1,303 primary, 22 refractory, and 37 relapse samples. One hundred sixty-four matched samples-127 primary and 37 relapse samples-were analyzed by using RNA sequencing. Results By using penalized lasso Cox proportional hazards regression, we identified 36 miRNAs the expression levels at diagnosis of which were highly associated with event-free survival. Combined expression of the 36 miRNAs was used to create a novel miRNA-based risk classification scheme (AMLmiR36). This new miRNA-based risk classifier identifies those patients who are at high risk (hazard ratio, 2.830; P ≤ .001) or low risk (hazard ratio, 0.323; P ≤ .001) of experiencing treatment failure, independent of conventional karyotype or mutation status. The performance of AMLmiR36 was independently assessed by using 878 patients from two different clinical trials (AAML0531 and AAML1031). Our analysis also revealed that miR-106a-363 was abundantly expressed in relapse and refractory samples, and several candidate targets of miR-106a-5p were involved in oxidative phosphorylation, a process that is suppressed in treatment-resistant leukemic cells. Conclusion To assess the utility of miRNAs for outcome prediction in patients with pediatric AML, we designed and validated a miRNA-based risk classification scheme. We also hypothesized that the abundant expression of miR-106a could increase treatment resistance via modulation of genes that are involved in oxidative phosphorylation.</p>

10.1200/JCO.2017.74.7451

J. Clin. Oncol.

29068783

CD33 Splicing Polymorphism Determines Gemtuzumab Ozogamicin Response in De Novo Acute Myeloid Leukemia: Report From Randomized Phase III Children's Oncology Group Trial AAML0531.

2017

JCO2016712513

2017 Jun 23

1527-7755

<p>Purpose Gemtuzumab ozogamicin (GO), a CD33-targeted immunoconjugate, is a re-emerging therapy for acute myeloid leukemia (AML). CD33 single nucleotide polymorphism rs12459419 C&gt;T in the splice enhancer region regulates the expression of an alternatively spliced CD33 isoform lacking exon2 (D2-CD33), thus eliminating the CD33 IgV domain, which is the antibody-binding site for GO, as well as diagnostic immunophenotypic panels. We aimed to determine the impact of the genotype of this splicing polymorphism in patients with AML treated with GO-containing chemotherapy. Patients and Methods CD33 splicing single nucleotide polymorphism was evaluated in newly diagnosed patients with AML randomly assigned to receive standard five-course chemotherapy alone (No-GO arm, n = 408) or chemotherapy with the addition of two doses of GO once during induction and once during intensification (GO arm, n = 408) as per the Children's Oncology Group AAML0531 trial. Results The rs12459419 genotype was CC in 415 patients (51%), CT in 316 patients (39%), and TT in 85 patients (10%), with a minor allele frequency of 30%. The T allele was significantly associated with higher levels of D2-CD33 transcript ( P &lt; 1.0E(-6)) and with lower diagnostic leukemic cell surface CD33 intensity ( P &lt; 1.0E(-6)). Patients with the CC genotype had significantly lower relapse risk in the GO arm than in the No-GO arm (26% v 49%; P &lt; .001). However, in patients with the CT or TT genotype, exposure to GO did not influence relapse risk (39% v 40%; P = .85). Disease-free survival was higher in patients with the CC genotype in the GO arm than in the No-GO arm (65% v 46%, respectively; P = .004), but this benefit of GO addition was not seen in patients with the CT or TT genotype. Conclusion Our results suggest that patients with the CC genotype for rs12459419 have a substantial response to GO, making this a potential biomarker for the selection of patients with a likelihood of significant response to GO.</p>

10.1200/JCO.2016.71.2513

J. Clin. Oncol.

28644774