Factors Associated with Progression to Overt Acute Myeloid Leukemia in NPM1-Mutated Myeloid Neoplasms with Less Than 10% Blasts

Introduction: The updated 5th World Health Organization (WHO) classification no longer restricts the diagnosis of acute myeloid leukemia (AML) with NPM1 mutation with blast percentage, while the International Consensus Classification (ICC) includes the criteria of 10% blasts, considering that blast percentage reflects biological heterogeneity, leading to confusions in diagnose and management ofNPM1-mutated myeloid neoplasms (MN) patients, especially those with less than 10% blasts. Therefore, we conducted this study to assess the clinical and molecular characteristics as well as the outcomes of NPM1-mutated MN with less than 10% blasts, to offer a comprehensive undestanding of these patients.

Methods: This retrospective study involved 42 newly diagnosed MN patients with NPM1 mutation and less than 10% blasts in bone marrow and peripheral blood at our center from August 30, 2016, to August 31, 2023, and were followed through December 30, 2023. Clinical and laboratory data at diagnosis were collected. Bone marrow samples at diagnosis were available for all subjects. In addition, 24 NPM1-mutated patients with 10-19% blasts from our center and 118 cases of de novo NPM1-mutated overt AML from the Beat AML progrom were obtained for comparision of demographic, cytogenetic and genomic features. In order to reveal a more precise clinical course of patients with < 10% blasts, we used Fine-Gray model（competing risk model）to evaluate risk factors associated with progression to overt AML, with death and allo hematopoietic stem cell transplantation (allo-HSCT) before progression as competing risks. Impact of allo-HSCT on OS was introducing allo-HSCT as a time-dependent covariate, represented using the Simon-Makuch method.

Results: Patients with NPM1 mutation shared similar clinical characteristics except for patients with < 10% blasts who had a younger median age compared to those with ≥20% blasts (49 vs 59 years, P< 0.001). There were no significant differences in individual genes between patients with blast < 10% and 10-19% blasts, except for counting mutations in the RAS pathway together (including NRAS, KRAS, PTPN11, NF1, CBL, P=0.035). Compared to overt NPM1-mutated AML, the most significant distinction was the low prevalence of FLT3-ITD mutations, whereas approximately 40% of overt AML had FLT3-ITD mutations. When subgrouping patients with <10% blasts according to ICC 2022, there was no difference in the frequency of individual genes between MDS and CMML group as well, except for more RAS passway mutations in CMML (P=0.04), similar to the aforementioned results. Notably, there were 8 MDS patients with BCORmutations, 2 of whom had multiple hits, and only 1 of the CMML patients had BCOR mutations, although statistical significance was not reached.

Four patients were lost to follow-up. Paients with FLT3-ITD mutation were not included in the analyses due to its definitive effect onsynergizing to drive rapid overt AML. In our cohort, 3 patients had FLT3-ITD mutation, 2 of them progressed to overt AML after 3, 5 months after diagnosis, respectively and 1 underwent allo-HSCT. Median follow-up was 50 months (95%CI，32.4-67.6 months). Multivariable analysis showed that VAF ≥ 45% (HR, 4.27 [95%CI, 1.21-15.1], P=0.024), monocyte count ≥ 0.38×10E+9/L (HR, 3.30 [95%CI, 1.09-10.1], P=0.035), and TET2 (HR, 1.88 [95%CI, 1.55-27.9], P=0.01) were associated with progression to overt AML in patients with <10% blasts. Among patients with 10-19% blasts, only DNMT3A mutation was associated with progression in univariable analysis. Allo-HSCT was significantly associated with improved OS in both <10% and 10-19 % blasts group.

Conclusion: The boundary of 10% blasts was not sufficient to distinguish the disease entity. Among patients with <10% blasts, patients with VAF ≥ 45%, monocyte count ≥ 0.38×10E+9/L and TET2 mutation are at high risk of progressing to overt AML. ICC criterion would cause clinicians underestimate the risk of NPM1-mutated MN with low blast percentage, and lead to death due to rapid progression and delayed follow-up and treatment. Our study supported the current WHO classifications to recognize NPM1-mutated MNs as a unique entity regardless of blast percentage.