<i>NPM1</i> mutation but not <i>RUNX1</i> mutation or multilineage dysplasia defines a prognostic subgroup within de novo acute myeloid leukemia lacking recurrent cytogenetic abnormalities in the revised 2016 WHO classification
Authors: Olga K. Weinberg, Christopher J. Gibson, Traci M. Blonquist, Donna Neuberg, Olga Pozdnyakova, Frank Kuo, Benjamin L. Ebert, Robert P. Hasserjian
Journal: American Journal of Hematology (2017)

Acute myeloid leukemia (AML) is a clinically and biologically heterogeneous disease, and several classification systems have been devised to refine prognostication and inform clinical decision-making. The World Health Organization (WHO) classification is widely used throughout the United States and internationally to classify AML in the clinical setting. The 4th edition classification was published in 2008 and a revised (2016) 4th edition classification has recently been proposed.1 This classification divides AML into groups with recurrent genetic abnormalities (AML-RGA), AML with myelodysplasia-related changes (AML-MRC), therapy-related myeloid AML, and AML not otherwise specified (AML-NOS).1 AML with mutated NPM1 (AML-NPM1) and with biallelic CEBPA mutation (AML-CEBPA), considered as provisional entities in the 2008 Classification, are now considered as distinct entities in the 2016 Classification; moreover, these mutations now supersede morphologic multilineage dysplasia.1 AML with RUNX1 mutation (AML-RUNX1) is a new provisional AML-RGA disease category.1 In the context of altered definitions in the revised WHO classification, the prognostic significance of the mutation-defined AML-RGA entities relative to AML-NOS and AML-MRC is uncertain.2-4 We assessed the ability of the revised 2016 WHO AML categories to predict prognosis within a cohort of patients with clinically-defined de novo AML lacking WHO-defined recurrent cytogenetic abnormalities. We identified cases of newly diagnosed de novo AML from the pathology archives of Brigham and Women's Hospital and Massachusetts General Hospital between 2009 and 2015. We excluded patients who had received prior cytotoxic therapy, carried a prior diagnosis of any myeloid neoplasm, or had WHO-defined recurrent cytogenetic abnormalities that would characterize the cases as AML-RGA or AML-MRC. All patients were treated with standard induction chemotherapy. Slides from each case were viewed in a blinded fashion by 3 hematopathologists (OW, RH and OP) to assign cases to the AML-MRC group according to the WHO classification criteria (at least 50% dysplastic cells in at least two lineages). We also quantified micromegakaryocytes, which have been found to have significant prognostic impact on de novo AML lacking WHO-defined cytogenetic abnormalities.5 For all patients meeting inclusion criteria, we performed targeted sequencing of 87 genes recurrently mutated in myeloid malignancies on either banked DNA samples or banked bone marrow aspirate samples as described previously.6 We classified variants as pathogenic driver mutations based on mutation type, position and frequency in publicly available SNP databases. Variants with variant allele fraction less than 0.02 were excluded. We compared clinical outcomes between patients based on their classification within the 2016 WHO scheme. Overall survival (OS) and event free survival (EFS) were defined as previously reported.5 We assessed EFS with Cox proportional hazards models, identifying SCT as a time-dependent variable; models were also adjusted for age and white blood count (WBC). We identified 137 patients who met inclusion criteria. The median age was 58.5 years, and most (112/137, 82%) patients had a normal karyotype. The most common cytogenetic abnormalities were +8 (six cases), +11 (four cases), and +13 (two cases); other trisomies were seen in four cases, chromosome losses in three cases, ring chromosome in one case, and balanced or unbalanced translocations in five cases. Of the 137 patients, 123 (90%) achieved complete remission (CR). 92 (67%) patients underwent stem cell transplant (SCT), 66 in first CR and 26 after experiencing relapse. According to the 2016 WHO classification, our cohort of patients included 41 AML-NOS (30%), 16 AML-MRC (12%), 56 AML-NPM1 (41%), 22 AML-RUNX1 (16%) and 2 AML-CEBPA (1%), compared to 55 AML-NOS (40%), 42 AML-MRC (31%), 37 AML-NPM1 (27%) and 3 AML-CEPBA (2%) based on 2008 WHO criteria. Most of the AML-RUNX1 cases were classified in the 2008 WHO as AML-NOS (17/22), with 5 AML-MRC. One case classified as AML-RUNX1 had both RUNX1 and NPM1 mutations, but was classified as AML-RUNX1 based on a higher RUNX1 mutant allele fraction (43%) versus NPM1 (29%). Given the small number of cases in the AML-CEBPA category, it was excluded from further analysis. When adjusting for SCT, age, and WBC, only the AML-NPM1 subgroup showed a significant difference in EFS compared to AML-NOS (p = 0.02, Table 1). Neither AML-MRC as defined by 2016 WHO classification nor AML-RUNX1 had significantly different EFS compared to AML-NOS. However, using an altered definition of AML-MRC based on the presence of ≥50% micromegakaryocytes in the absence of NPM1 or RUNX1 mutations, the micromegakaryocyte-defined AML-MRC group was associated with significantly shorter EFS (p = 0.004) as compared to AML-NOS (which comprised the remaining cases lacking NPM1 or RUNX1 mutations, Table 1). The 2016 revision to the WHO classification for AML incorporates morphologic, cytogenetic, and genetic criteria for risk stratification and is based on the consensus of expert hematopathologists and clinicians. Within our patient cohort, according to the 2016 WHO criteria, AML-NPM1 comprised a higher proportion of cases (41%) than using the 2008 criteria (27%), due to the allowance of multilineage dysplasia in the revised classification. Additionally, a significant proportion of patients in this cohort (16%) were diagnosed as AML-RUNX1. Conversely, compared to the 2008 WHO Classification, fewer cases were classified as AML-MRC based on MRC-defining morphology (12% in 2016 as compared to 31% in the 2008 WHO classification). We found that, while the 2016 WHO provisional entity of AML-RUNX1 had significantly shorter EFS compared to AML-NPM1 (P = .006, data not shown), there was no significant difference in EFS between AML-RUNX1 and AML-NOS. We also found that, while the morphology-based AML-MRC category was not significantly associated with EFS in this cohort, the specific morphologic finding of ≥50% micromegakaryocytes conferred an adverse prognosis. This suggests that a more restricted definition of myelodyplasia-related morphology based on micromegakaryocytes may result in more relevant morphology-based risk stratification within de novo AML lacking WHO-defined cytogenetic abnormalities or NPM1 or RUNX1 mutations. In summary, our findings support the prognostic relevance of the new 2016 WHO definition of AML-NPM1 that allows the presence of multilineage dysplasia. There was no prognostic impact of AML-MRC in this patient cohort; however, our results suggest that micromegakaryocytes may be a better marker of MDS-related ontogeny than WHO-defined multilineage dysplasia.