Related Experiment Video
Updated: Apr 5, 2026
Comet Assay to Quantify DNA Damage in FLT3 Mutant-expressing 32D Cells after Exposure to Type I and Type II FLT3 Inhibitors
Published on: October 17, 2025
Proteasome inhibitors induce FLT3-ITD degradation through autophagy in AML cells
Clément Larrue1, Estelle Saland1, Héléna Boutzen1
1Cancer Research Center of Toulouse, Unité Mixte de Recherche (UMR)1037 INSERM, ERL5294 Centre National de la Recherche Scientifique (CNRS), Toulouse, France; Université Toulouse III Paul Sabatier, Toulouse, France;
Abstract:
Internal tandem duplication of the Fms-like tyrosine kinase-3 receptor (FLT3) internal tandem duplication (ITD) is found in 30% of acute myeloid leukemia (AML) and is associated with a poor outcome. In addition to tyrosine kinase inhibitors, therapeutic strategies that modulate the expression of FLT3-ITD are also promising. We show that AML samples bearing FLT3-ITD mutations are more sensitive to proteasome inhibitors than wild-type samples and this sensitivity is strongly correlated with a higher FLT3-ITD allelic burden. Using pharmacologic inhibitors of autophagy, specific downregulation of key autophagy proteins including Vps34, autophagy gene (Atg)5, Atg12, Atg13, biochemical, and microscopy studies, we demonstrated that proteasome inhibitors induced cytotoxic autophagy in AML cells. FLT3-ITD molecules were detectable within autophagosomes after bortezomib treatment indicating that autophagy induction was responsible for the early degradation of FLT3-ITD, which preceded the inhibition of mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK), PI3K/AKT, and STAT5 pathways, and subsequent activation of cell death. Moreover, proteasome inhibitors overcome resistance to quizartinib induced by mutations in the kinase domain of FLT3, suggesting that these compounds may prevent the emergence of mutant clones arising from tyrosine kinase inhibitor treatments. In xenograft mice models, bortezomib stimulated the conversion of LC3-I to LC3-II, indicating induction of autophagy in vivo, downregulated FLT3-ITD protein expression and improved overall survival. Therefore, selecting patients according to FLT3-ITD mutations could be a new way to detect a significant clinical activity of proteasome inhibitors in AML patients.
Insights
Proteasome inhibitors induce autophagy and degrade FLT3-ITD in acute myeloid leukemia (AML) cells, offering a new therapeutic strategy. This approach shows promise in overcoming resistance to tyrosine kinase inhibitors and improving patient survival.
Area of Science:
- Hematology
- Molecular Biology
- Cancer Research
Background:
- Internal tandem duplication (ITD) of the Fms-like tyrosine kinase-3 receptor (FLT3) occurs in 30% of acute myeloid leukemia (AML) cases, correlating with poor prognosis.
- Therapeutic strategies targeting FLT3-ITD, beyond tyrosine kinase inhibitors, are needed.
Purpose of the Study:
- To investigate the efficacy of proteasome inhibitors in AML with FLT3-ITD mutations.
- To elucidate the mechanism by which proteasome inhibitors exert their effects in FLT3-ITD positive AML.
Main Methods:
- Treatment of AML samples and cell lines with proteasome inhibitors (e.g., bortezomib).
- Assessment of autophagy induction via downregulation of autophagy proteins (Vps34, Atg5, Atg12, Atg13) and LC3-I/II conversion.
- Detection of FLT3-ITD within autophagosomes and analysis of downstream signaling pathways (MAPK/ERK, PI3K/AKT, STAT5).
- Evaluation of proteasome inhibitors in overcoming quizartinib resistance and in xenograft mouse models.
Main Results:
- AML samples with FLT3-ITD mutations demonstrated increased sensitivity to proteasome inhibitors, correlated with higher FLT3-ITD allelic burden.
- Proteasome inhibitors induced cytotoxic autophagy in AML cells, leading to the degradation of FLT3-ITD.
- FLT3-ITD degradation preceded the inhibition of key signaling pathways and subsequent cell death.
- Proteasome inhibitors overcame resistance to quizartinib caused by FLT3 kinase domain mutations.
- In vivo studies showed bortezomib induced autophagy, downregulated FLT3-ITD, and improved survival in xenograft models.
Conclusions:
- Proteasome inhibitors represent a promising therapeutic strategy for FLT3-ITD positive AML by inducing autophagic degradation of FLT3-ITD.
- This approach may overcome resistance to existing tyrosine kinase inhibitors and prevent the emergence of resistant clones.
- Patient selection based on FLT3-ITD mutations could identify individuals likely to benefit from proteasome inhibitor therapy.
Related Concept Videos
01:13The Proteasome
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. This involves participation of a series of enzymes including— E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
02:18The Proteasome
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. A series of enzymes carry out the ubiquitination of the target proteins - E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
02:58Regulated Protein Degradation
Protein degradation plays two important roles in the cells. It helps to protect cells from misfolded or damaged proteins before they lead to a...
01:27Autophagy
An autophagic pathway consists of a series of signaling events activated in response to diverse stress and physiological conditions such as food deprivation,...
01:18Autophagic Cell Death
Autophagy and Apoptosis
Autophagy can activate apoptosis. In normal conditions, the autophagy activating protein Beclin-1 and...
01:31The Intrinsic Apoptotic Pathway