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.

PURPOSE: Optimized strategies for risk classification are essential to tailor therapy for patients with biologically distinctive disease. Risk classification in pediatric acute myeloid leukemia (pAML) relies on detection of translocations and gene mutations. Long noncoding RNA (lncRNA) transcripts have been shown to associate with and mediate malignant phenotypes in acute myeloid leukemia (AML) but have not been comprehensively evaluated in pAML.

METHODS: To identify lncRNA transcripts associated with outcomes, we evaluated the annotated lncRNA landscape by transcript sequencing of 1,298 pediatric and 96 adult AML specimens. Upregulated lncRNAs identified in the pAML training set were used to establish a regularized Cox regression model of event-free survival (EFS), yielding a 37 lncRNA signature (lncScore). Discretized lncScores were correlated with initial and postinduction treatment outcomes using Cox proportional hazards models in validation sets. Predictive model performance was compared with standard stratification methods by concordance analysis.

RESULTS: Training set cases with positive lncScores had 5-year EFS and overall survival rates of 26.7% and 42.7%, respectively, compared with 56.9% and 76.3% with negative lncScores (hazard ratio, 2.48 and 3.16; < .001). Pediatric validation cohorts and an adult AML group yielded comparable results in magnitude and significance. lncScore remained independently prognostic in multivariable models, including key factors used in preinduction and postinduction risk stratification. Subgroup analysis suggested that lncScores provide additional outcome information in heterogeneous subgroups currently classified as indeterminate risk. Concordance analysis showed that lncScore adds to overall classification accuracy with at least comparable predictive performance to current stratification methods that rely on multiple assays.

CONCLUSION: Inclusion of the lncScore enhances predictive power of traditional cytogenetic and mutation-defined stratification in pAML with potential, as a single assay, to replace these complex stratification schemes with comparable predictive accuracy.

Risk stratification for acute myeloid leukemia (AML) uses molecular and cytogenetic abnormalities identified at diagnosis. Response to therapy informs risk, and morphology continues to be used more frequently than flow cytometry. Herein, the largest cohort of pediatric patients prospectively assessed for measurable residual disease (MRD) by flow cytometry (N = 784) is reported. The "difference from normal" (ΔN) technique was applied: 31% of all patients tested positive (AML range, 0.02% to 91%) after the first course of treatment on Children's Oncology Group study AAML0531. Detection of MRD following initial chemotherapy proved the strongest predicator of overall survival (OS) in univariable and multivariable analyses, and was predictive of relapse risk, disease-free survival, and treatment-related mortality. Clearance of MRD after a second round of chemotherapy did not improve survival. The morphologic definition of persistent disease (>15% AML) failed 27% of the time; those identified as MRD- had superior outcomes. Similarly, for patients not achieving morphologic remission (>5% blasts), 36% of patients were MRD- and had favorable outcomes compared with those who were MRD+ (P < .001); hence an increase in myeloid progenitor cells can be favorable when ΔN classifies them as phenotypically normal. Furthermore, ΔN reclassified 20% of patients in morphologic remission as having detectable MRD with comparable poor outcomes. Retrospective analysis using the relapse phenotype as a template demonstrated that 96% of MRD- patients had <0.02% of the relapse immunophenotype in their end of induction 1 marrow. Thus, the detection of abnormal myeloid progenitor cells by ΔN is both specific and sensitive, with a high predictive signal identifiable early in treatment. This trial was registered at www.clinicaltrials.gov as #NCT00372593.

The addition of the proteasome inhibitor (PI) bortezomib to standard chemotherapy (ADE: cytarabine [Ara-C], daunorubicin and etoposide,) did not improve overall outcome in the Children's Oncology Group AAML1031 phase 3 randomized clinical trial (AAML1031). Bortezomib prevents protein degradation, including RelA via the intracellular NF-kB pathway. In this study, we hypothesized that subgroups of pediatric AML patients benefitting from standard therapy plus bortezomib (ADEB) could be identified based on pre-treatment RelA expression and phosphorylation status. RelA-total and phosphorylation at serine 536 (RelA-pSer ) levels were measured in 483 patient samples using reverse phase protein array technology. In ADEB-treated patients, low-RelA-pSer was favorably prognostic when compared to high-RelA-pSer (3-yr overall survival (OS): 81% vs. 68%, p = 0.032; relapse risk (RR): 30% vs. 49%, p = 0.004). RR in low-RelA-pSer patients significantly decreased in ADEB compared to ADE (RR: 30% vs. 44%, p = 0.035). Correlation between RelA-pSer and 295 other assayed proteins identified a strong correlation with HSF1-pSer , another protein previously identified as modifying ADEB response. The combination of low-RelA-pSer and low-HSF1-pSer was a significant predictor of ADEB response (3-yr OS: 86% vs. 67%, p = 0.013). Thus, bortezomib may improve clinical outcome in a subgroup of AML patients identified by low-RelA-pSer and low-HSF1-pSer . This article is protected by copyright. All rights reserved.

<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>

<p>Adolescent and young adult (AYA) patients with acute leukemia (AL) have inferior outcomes in comparison to younger patients, and are more likely to develop acute and chronic GVHD than younger children following HLA matched sibling donor stem cell transplant (SCT). We compared the incidence of grade II-IV acute GVHD, chronic GVHD, and survival in AYA (age 13-21 years) to younger children (age 2-12 years) who received an unrelated donor SCT for acute leukemia on Children's Oncology Group trials between 2004-2017. One hundred and eighty-eight children and young adults ages 2-21 years underwent URD SCT. Sixty-three percent were aged 2-12 and 37% were age 13-21. Older age was a risk factor for grade II-IV acute GVHD in multivariate analysis with a hazard ratio (HR) of 1.95 [95% confidence interval (CI) 1.23-3.10], but not for chronic GVHD, HR 1.25 [95% CI 0.57-2.71]. Younger patients relapsed more often (34.5 ± 4.4% vs. 22.8 ± 4.0%, p = 0.032), but their Event-Free Survival (42.6 ± 4.7% vs. 51.8 ± 6.1%, p = 0.18) and Overall Survival at 5 years (48.5 ± 4.9% vs. 51.5 ± 6.4%, p = 0.56) were not different than AYA patients. AYA patients who receive an URD SCT for acute leukemia are significantly more likely to develop grade II-IV acute GVHD, though survival is similar.</p>

<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>

<p><strong>BACKGROUND: </strong>High-risk pediatric acute myeloid leukemia confers a poor prognosis, and alternative strategies are needed to improve outcomes. We hypothesized that intensifying induction on the AAML1031 clinical trial would improve outcomes compared to the predecessor trial AAML0531.</p>

<p><strong>METHODS: </strong>Patients on AAML0531 received cytarabine (1600&nbsp;mg/m )/daunorubicin (150&nbsp;mg/m )/etoposide (ADE) for induction II and patients on AAML1031 received mitoxantrone (48&nbsp;mg/m )/cytarabine (8000&nbsp;mg/m ) (MA). Stem cell transplant (SCT) conditioning included busulfan/cyclophosphamide on AAML0531, whereas AAML1031 used busulfan/fludarabine and liberalized donor eligibility. Patients were included in this analysis if they met high-risk criteria common to the two trials by cytogenics or poor disease response after induction I ADE.</p>

<p><strong>RESULTS: </strong>MA provided no benefit over ADE at: induction II response (complete response [CR]: 64% vs. 62%, p&nbsp;=&nbsp;.87; measurable residual disease [MRD]+: 57% vs. 46%, p&nbsp;=&nbsp;.34); or intensification I response (CR: 79% vs. 94%, p&nbsp;=&nbsp;.27; MRD+: 27% vs. 20%, p&nbsp;=&nbsp;1.0). When considered with altered SCT approach, MA did not improve 5-year disease-free survival (24% ± 9% vs. 18% ± 15%, p&nbsp;=&nbsp;.63) or 5-year overall survival (35% ± 10% vs. 38% ± 18%, p&nbsp;=&nbsp;.66). MA was associated with slower neutrophil recovery (median 34 vs. 27&nbsp;days, p&nbsp;=&nbsp;.007) and platelet recovery (median 29 vs. 24.5&nbsp;days, p&nbsp;=&nbsp;.04) and longer hospital stay (32 vs. 28&nbsp;days, p&nbsp;=&nbsp;.01) during induction II.</p>

<p><strong>CONCLUSION: </strong>Intensification of induction II did not improve treatment response or survival, but did increase toxicity and resource utilization. Alternative strategies are urgently needed to improve outcomes for pediatric patients with high-risk acute myeloid leukemia (trials registered at clinicaltrials.gov NCT01371981, NCT00372593).</p>

<p>Post-relapse therapy remains critical for survival in children with acute myeloid leukemia (AML). We evaluated survival, response and prognostic variables following relapse in independent cooperative group studies conducted by COG and the population-based AML-BFM study group. BFM included 197 patients who relapsed after closure of the last I-BFM relapse trial until 2017, while COG included 852 patients who relapsed on the last Phase 3 trials (AAML0531, AAML1031). Overall survival at 5 years (OS) was 42 ± 4% (BFM) and 35 ± 2% (COG). Initial high-risk features (BFM 32 ± 6%, COG 26 ± 4%) and short time to relapse (BFM 29 ± 4%, COG 25 ± 2%) predicted diminished survival. In the BFM dataset, there was no difference in OS for patients who had a complete remission with full hematopoietic recovery (CR) following post-relapse re-induction compared to those with partial neutrophil and platelet recovery (CRp and CRi) only (52 ± 7% vs. 63 ± 10%, = 0.39). Among 90 patients alive at last follow-up, 87 had received a post-relapse hematopoietic stem cell transplant (HSCT). OS for patients with post-relapse HSCT was 54 ± 4%. In conclusion, initial high-risk features and early relapse remain prognostic. Response assessment with full hematopoietic recovery following initial relapse therapy does not predict survival. These data indicate the need for post-relapse risk stratification in future studies of relapse therapies.</p>

<p><strong>PURPOSE: </strong>We investigated the impact of the CD33-targeted agent gemtuzumab ozogamicin (GO) on survival in pediatric patients with -rearranged (-r) acute myeloid leukemia (AML) enrolled in the Children's Oncology Group trial AAML0531 (NCT01407757).</p>

<p><strong>METHODS: </strong>Patients with -r AML were identified and clinical characteristics described. Five-year overall survival (OS), event-free survival (EFS), disease-free survival (DFS), and relapse risk (RR) were determined overall and for higher-risk versus not high-risk translocation partners. GO's impact on response was determined and outcomes based on consolidation approach (hematopoietic stem cell transplant [HSCT] chemotherapy) described.</p>

<p><strong>RESULTS: </strong>Two hundred fifteen (21%) of 1,022 patients enrolled had -r AML. Five-year EFS and OS from study entry were 38% and 58%, respectively. EFS was superior with GO treatment (EFS 48% with GO 29% without, = .003), although OS was comparable (63% 53%, = .054). For patients with -r AML who achieved complete remission, GO was associated with lower RR (40% GO 66% patients who did not receive GO [No-GO], = .001) and improved 5-year DFS (GO 57% No-GO 33%, = .002). GO benefit was observed in both higher-risk and not high-risk -r subsets. For patients who underwent HSCT, prior GO exposure was associated with decreased relapse (5-year RR: 28% GO and HSCT 73% No-GO and HSCT, = .006). In multivariable analysis, GO was independently associated with improved EFS, improved DFS, and reduced RR.</p>

<p><strong>CONCLUSION: </strong>GO added to conventional chemotherapy improved outcomes for -r AML; consolidation with HSCT may further enhance outcomes. Future clinical trials should study CD33-targeted agents in combination with HSCT for pediatric r AML.</p>