Minimal Residual Disease Risk-Stratification in Pediatric Mixed Phenotype Acute Leukemia: Results of a Multi-Center Cohort Study
Authors: Oberley Matthew J. , Raikar Sunil S. , Malvar Jemily , Wertheim Gerald , Seif Alix E. , Guinipero Terri , Sposto Richard , Rabin Karen R. , Punia Jyotinder N. , Schore Reuven J. , Luca Dragos C. , Woods William G. , O'Gorman Maurice R.G. , Orgel Etan 
Published: October 2019

Abstract
BACKGROUND: Mixed phenotype acute leukemia (MPAL) is a rare form of leukemia in children. Current evidence supports the use of acute lymphoblastic leukemia (ALL)-directed regimens as initial therapy for MPAL; Children's Oncology Group (COG) ALL regimens are commonly used for pediatric ALL in the United States. Data for the predictive value of minimal residual disease (MRD) to risk-stratify therapy for pediatric MPAL is sparse overall and currently unknown in the context of COG ALL regimens. The primary objective for this study was to therefore examine the predictive value of MRD for event-free and overall survival (EFS, OS) in a centrally-reviewed, strictly-defined pediatric MPAL cohort treated according to COG ALL regimens.
METHODS: A retrospective cohort of pediatric MPAL treated from 2008-2016 was assembled from six institutions in the United States. All sites submitted primary diagnostic flow cytometry for central review. Two independent hematopathologists blinded to clinical outcomes confirmed the diagnosis of MPAL according to the WHO2008 and/or WHO2016 classification and assigned a MPAL phenotype. MRD was assessed by multi-parameter flow cytometry ("negative" defined as <0.01%). Endpoints of interest included presenting features of MPAL, MRD levels at end of induction and consolidation (EOI, EOC), role of hematopoietic stem cell transplantation (HSCT) in complete remission (CR), EFS, and OS. Kaplan-Meier analyses were performed stratified by predictors of interest. Stepwise multivariable Cox proportional hazard models inclusive of MRD and candidate predictors were used to evaluate EFS and OS for patients with complete data. Analyses were repeated in the subset of patients with "neutral" cytogenetics (i.e. no favorable [double trisomy, ETV6-RUNX1] or adverse [hypodiploid, BCR-ABL1, KMT2Ar, Ph-like, iAMP21, FLT3-ITD] features). All tests were 2-sided with significance set at p <0.05.
RESULTS: Of the 112 cases diagnosed by institutions as MPAL, 94 (84%) fulfilled strict WHO criteria. The majority of MPAL patients were B/Myeloid (89%), with a single blast population (87%), and without hyperleukocytosis (WBC≥50K/uL) or central nervous system involvement at presentation (Table 1). Within the cohort, 85/94 were treated with a COG ALL regimen with EOI MRD available (1/85 suffered an induction death prior to MRD assessment). ALL induction therapy resulted in a MRD-negative CR in 72% (61/85); three MRD-negative patients nonetheless converted to non-ALL therapy. Of those EOI MRD+ (≥0.01%), 12 of 14 who continued with COG ALL therapy achieved an EOC MRD-negative CR (i.e. 70/85 [82%] were MRD-negative by EOC). Induction failure (IF) occurred in 7% (6/85) and was defined as disease progression prior to EOI (n=2) or by EOI MRD ≥5% (n=4). The two patients with early disease progression were successfully "salvaged" with AML therapy (<21 days from initial diagnosis) and achieved a MRD-negative CR following one cycle. Patients achieving MRD-negative remissions at EOI and/or EOC with ALL chemotherapy had excellent EFS and OS (Figure 1). Multivariable analysis confirmed the predictive value of MRD at EOI (hazard ratio ±SE for EFS and OS = 3.77±1.67 and 3.54±2.09; p=0.032 and p=0.024 respectively). Only a few patients receiving ALL therapy proceeded to HSCT in CR1, all were alive at time of reporting. Use of HSCT was not associated with improved EFS or OS (p>0.50 for both); five-year EFS and OS for ALL chemotherapy without HSCT (n=73) were 78±6% and 90±4%, respectively. The presence of multiple blast populations of distinct lineages at diagnosis was similarly rare and was potentially associated with worse OS on multivariable analysis (p=0.024). No other candidate predictors were associated with EFS or OS. No difference in the predictive value of MRD was present when analyses were limited to patients with "neutral" cytogenetics.
CONCLUSION: In a retrospective analysis of pediatric MPAL, the majority of patients treated with ALL chemotherapy achieved a MRD-negative CR by EOC (~week 12 of therapy); overall survival in this group was excellent. Further prospective validation of MRD is essential to refine risk-stratified therapy for pediatric MPAL. Optimal salvage for those who fail to achieve remission with ALL chemotherapy is unknown and requires further study.

Disclosures
O'Gorman: Becton Dickinson: Consultancy.