Evaluating Impact of Insertion Length on Clinical Outcomes in FLT3-ITD Mutated AML: A Single Center Retrospective Analysis
Authors: Rauf Asad , Nurse Daniel P. , El-Asmar Jessica , Bawwab Ameed , Abu-Farsakh Yomna , Albliwi Moath , Hanna John , Abuamsha Hasan , Batah Heya , Zabor Emily C. , Bedelu Yohanna B. , Chen Mark Jinan , Nakitandwe Joy , Elbaz Younes Ismail , Bosler David S. , Jain Akriti G. , Molina John C. , Balderman Sophia R. , Singh Abhay , Gerds Aaron T. , Mukherjee Sudipto , Advani Anjali S. , Carraway Hetty E. , Mustafa Ali Moaath K. 
Published: December 2024

Background:
Nearly one-third of patients with acute myeloid leukemia (AML) present with mutations in the fms-like tyrosine kinase 3 (FLT3) gene, generally resulting in constitutive activation of downstream proliferative and prosurvival pathways. The most common FLT3 mutations among AML patients are internal tandem duplication (ITD) mutations. Although FLT3-ITD mutations are heterogeneous in size and location, they are frequently associated with higher allelic burdens and poor prognosis. There are currently conflicting data about how the length of the FLT3-ITD mutations affects patient outcomes or clinical decision-making. This study aims to determine how ITD insertion length impacts outcomes in patients with FLT3-ITD mutated AML.
Methods:
We conducted a single-center retrospective cohort study to compare overall survival (OS), event-free survival (EFS), clinical response rate, and cytogenetic response rate for FLT3-ITD mutated AML in adult patients (≥18 years) with various ITD insertion lengths. Three cohorts were stratified by mutation lengths of 0-33 base pairs (BP) (1-33 percentile), 34-66 BP (34-67 percentile), and &gt;66 BP (≥68 percentile). Insertion length was determined by polymerase chain reaction (PCR) or, for patients without PCR results, next-generation sequencing (NGS). Baseline variables were collected and included age, gender, ethnicity, comorbidities, and ECOG performance status. Factors associated with response were analyzed using multivariable logistic regression, with variables included as supported by the number of events. Overall survival time was calculated from the date of diagnosis to the date of death or last follow-up, whichever came first. Event-free survival was calculated from the date of treatment start to the date of first refractory disease or death from any cause, and patients alive without refractory disease were censored at the last date their disease status was known. The Kaplan-Meier method estimated survival probabilities, and the log-rank test was used to test for differences. Cox regression was used for multivariable analysis.
Results:
Between 5/2015 and 12/2023, 730 patients were diagnosed with AML, and 101 patients tested positive for FLT3-ITD and were available for analysis at the time of diagnosis at the Cleveland Clinic. The number of patients with FLT3-ITD mutation lengths of 0-33 BP, 34-66 BP, and &gt; 66 BP were 34 (34%), 33 (33%), and 34 (34%), respectively. The median ages for the subgroups were 66 years for 0-33 BP, 62 years for 34-66 BP and 67 years for &gt;66 BP groups (P=0.7). There was no difference in other demographic baseline or cytogenetic/ molecular features (all P&gt;0.05). Moreover, the 3 subgroups had similar hematologic parameters at the time of presentation (i.e., BM blasts, platelets, white blood cells) (P&gt;0.05).
Regarding outcomes, the CCR rates were 55% for 0-33 BP, 52% for 34-66 BP, and 50% for &gt;66 BP groups (P=0.9). Similarly, using multivariable logistic regression, the odds of achieving CCR in the subgroups were not different (P=0.8). The median follow-up was 33.51 months (range 2.07 - 90.18). On multivariable Cox proportional hazards model, the risk of mortality was not different in the &gt;66 BP group (hazard ratio (HR): 1.09, 95%CI: 059-2.01) or 34-66 BP group (HR:1.75, 95%CI 0.96-3.18) when compared to the 0-33 BP group (P=0.2). Similarly, there was no difference in EFS in the &gt;66 BP group (HR: 1.11, 95%CI: 0.62-2.0) or 34-66 BP group (HR:1.6, 95%CI 0.9-2.86) when compared to the 0-33 BP group (P=0.3).
Conclusion:
In our single-center retrospective study, the length of FLT3-ITD mutation insertions did not have any impact on clinical response rates, EFS, OS, or cytogenetic response. It is unknown if ITD length impacts the cytotoxicity level of FLT3 inhibitors. Moreover, less is known if response outcomes to FLT3 inhibitors vary based on the location of ITD (i.e., transmembrane vs intracellular domains).