Prevalence of FLT3 gene mutation and its expression in Brazilian pediatric B-ALL patients: clinical implications

Abstract

Introduction: There is consistent evidence that FLT3 may be a driver gene in B-ALL and that selected cases may benefit from the use of FLT3 inhibitors. Our study was conducted to evaluate the frequency and types of FLT3 mutations in pediatric patients with B-ALL, the relative expression of this gene, and their influence on clinical evolution.

Methods: We evaluated 156 children with B-ALL treated between July 2018 and September 2023. Screening for FLT3 mutations was performed using RFLP and fragment analysis, while FLT3 expression was assessed by qPCR.

Results: FLT3-TKD and/or FLT3-JM-INDEL mutations were found in 8 patients (5.1%). We did not identify any ITD-type mutations. None of the patients with identified FLT3 mutations presented recurrent rearrangements in B-ALL or alterations in the IKZF1, PAX5, or ERG genes, suggesting that FLT3 mutation may serve as the driving mechanism for leukemia in these cases. Two (2/8) patients with FLT3 mutations experienced disease relapse. Although we did not observe FLT3 overexpression among patients with FLT3 mutations, FLT3 expression levels were higher in these patients compared to WT patients. Four FLT3-WT patients presented FLT3 overexpression, defined as RQ > 10. FLT3 mutations or overexpression were not associated with relapses or survival rates.

Discussion: Our findings do not support the inclusion of FLT3 as a routine marker in the risk stratification of B-ALL patients; nevertheless, FLT3 alterations may be relevant for guiding personalized treatment approaches in specific clinical contexts.

Keywords: child health; molecular biology; precision medicine; precursor B-cell lymphoblastic leukemia-lymphoma; tumor biomarkers.

Figure 1

FLT3 mutations and expression assessment in 156 children with B ALL diagnosis (RQ: relative quantification).

Figure 2

Clinical and molecular profile of the 156 patients included in the study. Interm. Risk: intermediary risk. Low temp: low risk temporary (GBTLI protocol). True LR: true low risk (GBTLI protocol). IR: intermediary risk. Extreme HR: extreme high risk. High hyperd: high hyplerdiploidy; Low hyperdip: Low hyperdiploidy. Hipodip: hypodiploidy. Strutural alt: structural alteration. mFLT3: FLT3 mutation. eFLT3: FLT3 expression. In red, in the left column, patients with FLT3 overexpression (RQ > 10) or mutation are highlighted.

Figure 3

Description of eight distinct FLT3 Gene Mutations in eight (8) patients with B-ALL (nine occurrences). FLT3-JM-INDEL mutations were identified in four patients. Five patients had mutations in the tyrosine kinase domain (FLT3-TKD). One patient exhibited two mutations: one in the juxtamembrane domain (§) and another in the tyrosine kinase domain (§). Two patients presented the p.Ile836del mutation, with one patient having this mutation exclusively and the other having it in conjunction with a FLT3-JM-INDEL (§).

Figure 4

Relapse occurrence in patients with FLT3 gene mutation (n = 8) and wild type—WT (n = 147). Statistically analyzed by Fisher's exact test.

Figure 5

Survival curves for B-ALL pediatric patients according to status of FLT3 mutation (A) (global survival) and (B) (event free survival). Statistically analyzed by Log-rank (Mantel-Cox) test.

Figure 6

FLT3 expression analysis in samples of children with B-cell ALL. (A) Frequence distribution of FLT3 expression (relative quantification—RQ) in the studied population (attention to the log scale). Data did not pass in Shapiro-Wilk normality test and showed skewness of +8.69. (B) Evaluation of MRD at mid-induction (D15 for patients treated with the adapted BFM ALLIC 2009 protocol and D19 for patients treated with the Brazilian GBTLI 2021 protocol) among patients overexpressing FLT3 (>10) and all the others (<10). Statistical analyses were performed with Mann Whitney test. (C) Evaluation of MRD at the end of induction (D78 for patients treated with the BFL ALLIC 2009 and D49 for the group treated with the GBTLI protocol) among patients overexpressing FLT3 (>10) and all the others (<10). Statistical analyses were performed with Mann Whitney test. (D) FLT3 expression between patients with (FLT3 mut) and without (FLT3 WT) mutations. Statistical analyses were performed with Mann Whitney test.

Figure 7

FLT3 expression levels analysis in relapsed patients vs. patients in remission. (A) Comparison between FLT3 expression values of samples from 103 patients initially diagnosed with B-cell ALL who did not experience relapse until last data assessment and FLT3 expression values in BM samples collected at the initial diagnosis from 8 patients who relapsed. Statistically analyzed by Mann Whitney test. (B) Comparison between FLT3 expression values of samples from 112 patients initially diagnosed with B-cell ALL and FLT3 expression values of BM samples collected at the time of relapse from 10 patients. Statistically analyzed by Mann Whitney test. (C) FLT3 expression levels of 8 samples from patients at the initial diagnosis and at the time of relapse. The patient with the highest RQ value (2.5) at diagnosis is the only one in the group with FLT3 mutation. Statistically analyzed by Wilcoxon matched-pairs signed rank test.

Figure 8

Survival curves for B ALL pediatric patients according to FLT3 expression status. (A) (Overal Survival) and (B) (Event free survival). Statistically analyzed by Log-rank (Mantel-Cox) test.