FGF2 from Marrow Microenvironment Promotes Resistance to FLT3 Inhibitors in Acute Myeloid Leukemia: Beyond the Abstract

Activation of FLT3 by internal tandem duplication (FLT3-ITD) occurs in ~20% of acute myeloid leukemia (AML) patients and is associated with a higher rate of relapse1,2. A number of FLT3 inhibitors have been tested in the clinic, and while most patients respond initially, the responses are not durable and resistance typically develops within months. For some FLT3 inhibitors, such as quizartinib (AC220), mutations of the activation loop (most commonly D835) leads to resistance3. However activation loop mutations only accounts for a minority of clinical resistance.

In addition, AML patients treated with novel FLT3 inhibitors such as crenolanib and gilteritinib, which are not affected by D835 mutations, still develop resistance over time, indicating that other mechanisms of resistance are important. 

Although potent FLT3 inhibitors rapidly clear leukemia cells from the peripheral blood, the response within the bone marrow occurs more slowly. This suggests that survival signals from the bone marrow microenvironment protect residual AML cells, allowing development of resistance over time. We screened a number of proteins from the microenvironment and discovered that fibroblast growth factor 2 (FGF2) protected FLT3-ITD AML cell lines and primary FLT3-ITD AML cells from the effects of quizartinib in vitro. Continuous culture of cell lines with FGF2 and quizartinib led to development of resistance after a few months, similar to patients treated with quizartinib. 

We investigated the mechanism resistance and found that early resistance occurred through FGF2 activation of the FGF receptor (FGFR1) leading to downstream activation of the MAPK pathway, which provides an accessory survival pathway during FLT3 inhibition. Interestingly, continued culture with FGF2 and quizartinib frequently selected for FLT3 resistance mutations and/or activating mutations of RAS over time. Once resistance mutations developed, FGF2 activation of FGFR1-MAPK pathway became redundant and was no longer required for growth. 

We then evaluated FGF2 expression in the bone marrow of patients treated with quizartinib. FGF2 is normally expressed in bone marrow stromal cells, but expression of FGF2 increased significantly during treatment. This increase in marrow FGF2 correlated with development of early resistance. In parallel
with our in vitro results, once resistance mutations developed, FGF2 signaling became redundant, and stromal FGF2 expression decreased again. 

Taken together, our data supports a two-step model of resistance in FLT3-ITD AML. Initial resistance is mediated by ligands from the marrow microenvironment, allowing time for acquisition of resistance mutations, or other mechanisms of resistance. Increased expression of FGF2 has been shown to be activated in stress hematopoiesis4,5 and we hypothesize that this stress response is hijacked by leukemia cells for survival. Targeting early survival pathways in the microenvironment, such as FGF2-FGFR activation, is likely to reduce residual AML, and improve the durability of response.

More broadly, activation of FGFR appears to be a conserved mechanism of resistance in many kinase-driven malignancies. FGFRs have recently been shown to drive resistance in lung cancer6, gastrointestinal stromal tumors7,8 and chronic myeloid leukemia9. This argues for combination strategies that not only target the oncogenic kinase, but also accessory pathways like FGFR that are frequently activated in resistance.

Written by: Elie Traer, MD PhD

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