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Sphingosine-1-phosphate receptor modulators resensitize FLT3-ITD acute myeloid leukemia cells with NRAS mutations to
Aditi Chatterjee1,2, Moaath K Mustafa Ali1,2, Christopher M Bailey3
1University of Maryland Greenebaum Comprehensive Cancer Center, University of Maryland School of Medicine, Baltimore, MD, USA.
Abstract:
FLT3 inhibitor efficacy in AML with FLT3-ITD is short-lived, frequently due to new mutations, most commonly in NRAS. Sphingosine kinase 1 (SPHK1), which phosphorylates sphingosine to generate sphingosine-1-phosphate (S1P), is upregulated and localized to the plasma membrane in RAS-mutated cells. We studied S1P and FLT3 co-targeting to overcome FLT3 inhibitor resistance in NRAS-mutated FLT3-ITD AML cells. NRAS-mutated FLT3-ITD AML cell lines and patient blasts were treated with FLT3 inhibitors and/or S1P receptor (S1PR) modulators. FLT3 inhibitor sensitivity was assessed by immunoblotting, cytotoxicity, apoptosis and colony formation. Co-treatment was also assessed in vivo in an orthotopic mouse model. Downstream RAS and SPHK1 effectors were measured by immunoblotting and qRT-PCR. The S1PR modulators fingolimod (FTY720) and mocravimod (KRP-203) resensitized FLT3-ITD-expressing MOLM-14 and MV4-11 human AML cells with G12D, G12S, Q61K or Q61H, but not G12C, and patient blasts with G13D, G13V or G12D NRAS mutations to FLT3 inhibitors. Moreover, FTY720 co-treatment resensitized G12D NRAS-mutated M14(R)701 cells to gilteritinib in vivo. Co-treatment inactivated ERK, transcriptionally downregulated SPHK1, and inactivated downstream AKT, p70 S6K and BAD, with inactivation abrogated by constitutive SPHK1 expression. The clinically applicable S1PR modulators fingolimod and mocravimod resensitize NRAS-mutated FLT3-ITD AML cells to FLT3 inhibitors, supporting potential clinical efficacy.
Insights
FLT3 inhibitors are often ineffective against acute myeloid leukemia with FLT3-ITD mutations due to NRAS mutations. Combining S1P receptor modulators with FLT3 inhibitors can overcome this resistance, offering new hope for AML treatment.
Area of Science:
- Oncology
- Molecular Biology
- Pharmacology
Background:
- FLT3-ITD mutations drive acute myeloid leukemia (AML) but resistance emerges due to secondary mutations, commonly in NRAS.
- Sphingosine kinase 1 (SPHK1) is upregulated in RAS-mutated cells, producing sphingosine-1-phosphate (S1P), which may contribute to FLT3 inhibitor resistance.
Purpose of the Study:
- To investigate the co-targeting of S1P and FLT3 pathways to overcome FLT3 inhibitor resistance in NRAS-mutated FLT3-ITD AML.
- To evaluate the efficacy of S1P receptor (S1PR) modulators in combination with FLT3 inhibitors in preclinical AML models.
Main Methods:
- NRAS-mutated FLT3-ITD AML cell lines and patient blasts were treated with FLT3 inhibitors and/or S1PR modulators (fingolimod, mocravimod).
- Sensitivity was assessed via immunoblotting, cytotoxicity, apoptosis, and colony formation assays.
- In vivo efficacy was evaluated in an orthotopic mouse model; downstream signaling was analyzed by immunoblotting and qRT-PCR.
Main Results:
- S1PR modulators fingolimod and mocravimod resensitized various NRAS-mutated FLT3-ITD AML cell lines and patient blasts to FLT3 inhibitors.
- Co-treatment with FTY720 demonstrated in vivo efficacy in a mouse model of NRAS-mutated AML.
- Combination therapy led to inactivation of ERK, transcriptional downregulation of SPHK1, and inactivation of downstream AKT, p70 S6K, and BAD.
Conclusions:
- Clinically applicable S1PR modulators fingolimod and mocravimod can resensitize NRAS-mutated FLT3-ITD AML cells to FLT3 inhibitors.
- This combination strategy shows potential for overcoming FLT3 inhibitor resistance in AML.
- Targeting S1P signaling alongside FLT3 inhibition offers a promising therapeutic approach for resistant AML.
