Prognostic Role of NGS-Based MRD Assessment in FLT3-TKD Mutated Patients with Acute Myeloid Leukemia
Authors: Arnhardt Isabell , Vonk Christian M. , Tsai Xavier Cheng-Hong , Thol Felicitas R. , Chou Wen-Chien , Hou Hsin-An , Gabdoulline Razif , Heidel Florian H. , Tien Hwei-Fang , Heuser Michael , Valk Peter J. M. 
Published: December 2024

Background
The prognostic value of measurable residual disease (MRD) monitoring for patients with acute myeloid leukemia (AML) is well established for multicolor flow-cytometry, as well as AML patients with NPM1 mutation and FLT3internal tandem duplication (ITD). Next-Generation Sequencing (NGS) provides highly sensitive and comprehensive detection of MRD. However, due to the broad mutational landscape, the prognostic value of many genes with low frequency remains unclear. 7-10% of adult AML patients harbor a FLT3tyrosine kinase domain (FLT3-TKD) mutation, while its prognostic impact as MRD marker is still undetermined. We conducted a meta-analysis of published MRD studies using next-generation sequencing (NGS) to evaluate the prognostic impact of NGS-MRD in patients with FLT3-TKD mutation.
Methods
Published NGS-MRD data (Thol et al. Blood 2018; 132 (16): 1703-1713, Tsai et al. Blood Adv2021; 5 (10): 2456-2466, Jongen-Lavrencic et al. NEJM 2018; 378:1189-1199) were collected with pseudonymized, individual patient data. Patients in complete remission (CR) or complete remission with incomplete hematologic recovery (CRi) after intensive induction chemotherapy with available baseline clinical and genetic, treatment, response, and MRD data after two cycles of induction chemotherapy were included. The MRD status was determined by error-corrected targeted NGS or with the Illumina TruSight myeloid sequencing panel. MRD positivity was defined as two standard deviations above the background mean or variants above the background noise determined using outlier testing. The statistical analyses were performed using statistical software environment R, and statistical software package SPSS 27.0.
Results
From the three studies, 837 adult AML patients fulfilled the inclusion criteria. 90% were diagnosed with de novo AML, 407 (49%) were female, and the median age was 47.9 years (18-84). According to the ELN 2017 risk classification, 41.6% had favorable risk. Ninety-two patients (11%) carried a FLT3-TKD mutation at diagnosis, of whom 47 (51%) had co-mutated NPM1. Six of 92 FLT3-TKD AML patients (7%) were MRD positive for FLT3-TKD after two cycles of chemotherapy. The mean variant allele frequency (VAF) for FLT3-TKD positive patients was 0.18% (range: 0.009-0.75%).
FLT3-TKD MRD positive patients had similar clinical and genetic characteristics compared to MRD negative patients. However, the ELN 2017 risk classification differed between MRD negative and positive patients with 58 (67%) vs 0 (0%) classified as favorable risk, and 15 (17%) vs 4 (67%) classified as adverse risk. Only two patients in the MRD negative group received the FLT3 inhibitor midostaurin combined with chemotherapy. Three MRD positive patients (50%) underwent allogeneic hematopoietic cell transplantation (alloHCT) compared to 23 (27%) MRD negative patients.
The median follow up was 114.8 months (range 4-221 months). There was no significant difference in OS (HR: 1.85, 95% Cl: 0.65-5.24), RFS (HR: 1.17, 95% Cl: 0.36-3.82), and CIR (HR: 1.45, 95% Cl: 0.44-4.78) between FLT3-TKD MRD positive and negative patients.
We next evaluated MRD of these FLT3-TKD mutated patients based on non-DTA (DNMT3A, TET2, and ASXL1) mutations present at diagnosis evaluated after two cycles of chemotherapy. 32 (35%) patients were MRD positive for at least one marker. The mean VAF of non-DTA MRD was 0.37% (range 0.03-11.77%). Most frequently NPM1, IDH1, RUNX1, and SF3B1 mutations persisted in CR. MRD positive patients had similar clinical and genetic characteristics compared to MRD negative patients except that 12 (38%) MRD positive patients were considered favorable risk compared to 46 (77%) MRD negative patients. Comparing the prognostic impact of MRD using non-DTA gene mutations showed similar OS (HR: 1.6, 95% Cl: 0.85-3.02), RFS (HR: 1.24, 95% Cl: 0.65-2.38) and CIR (HR: 1.66, 95% Cl: 0.81-3.39) between patients with and without molecular MRD.
Conclusions
After two cycles of induction chemotherapy FLT3-TKD and non-DTA MRD remained detectable in 7% and 35%, respectively. NGS-MRD was not prognostic in this cohort of FLT3-TKD mutated AML patients.
Shared first authors: Isabell Arnhardt, Christian M Vonk
Shared senior authorship: Michael Heuser, Peter J.M. Valk