Contributions
Abstract: S817
Type: Oral Presentation
Presentation during EHA23: On Saturday, June 16, 2018 from 12:30 - 12:45
Location: Room A4
Background
Gilteritinib is a potent, selective FLT3 inhibitor with substantial single-agent clinical activity in relapsed/refractory (R/R) AML patients with FLT3 internal tandem duplication (ITD) and/or FLT3-D835 tyrosine kinase domain mutations (Perl AE, et al Lancet Oncol 2017). However, gilteritinib monotherapy is not curative and, like other FLT3 inhibitors, our data suggest acquired sequence mutations eventually arise and contribute to drug resistance (McMahon CM, et al, ASH 2017 abstract #295). In some cases, FLT3-wild type leukemic populations are selected during gilteritinib therapy and establish clonal dominance at clinical progression; in others, the original FLT3 mutation persists but is accompanied at progression by new mutations, often N-RAS or FLT3-F691L. Higher gilteritinib doses might combat resistance from FLT3-F691L, but combinatorial strategies are likely needed to circumvent other mechanisms of resistance and optimize response quality and duration.
Aims
We sought to describe and clarify mechanisms of secondary resistance to gilteritinib therapy.
Methods
Adults with R/R FLT3-mutated AML who received at least 80 mg/day of gilteritinib on a phase 1/2 study (NCT02014558) at 3 institutions provided leukemia samples at baseline, scheduled study time points, and at study withdrawal; all clinical molecular and cytogenetic data were reviewed. Baseline and progression samples were tested by a 66-gene next generation sequencing (NGS) panel; a subset with treatment-emergent mutations at disease progression were analyzed by a 24-gene single-cell NGS technique (Tapestri). Dual FLT3-ITD+/NRAS+ cell lines (MOLM14-NRAS) were generated through long-term culture of MOLM14 cells in low concentrations of the FLT3 inhibitor quizartinib. Effects of gilteritinib and/or the MEK inhibitor trametinib on the in vitro growth and apoptosis of MOLM14-NRAS cells were studied.
Results
29 subjects (median age 62) have complete, evaluable cytogenetic and/or NGS data; an additional 7 have data currently being analyzed. At study enrollment 26/29 (89.7%) had a FLT3-ITD mutation, including 5 (17.2%) with both FLT3-ITD and FLT3-D835 mutations. Three subjects (10.3%) had a FLT3-D835 mutation only. 23/29 progressed during gilteritinib therapy, 5 were withdrawn for transplant or toxicity, and one remains on gilteritinib in remission. 10/21 subjects (47.6%) with serial cytogenetic data developed new cytogenetic abnormalities during therapy, including a new BCR-ABL1 fusion at progression in 1 subject. Cytogenetic evolution did not always correlate with disease progression. New gene mutations developed on gilteritinib in 12/27 subjects (44.4%) with available NGS data, including NRAS (n=7), KRAS (n=2), FLT3-F691L (n=3), IDH2 (n=1), PTPN11 (n=2), TBL1XR1 (n=1), and CBL (n=1). Single-cell NGS confirmed that treatment-emergent RAS mutations indeed occur in FLT3-mutated cells. MOLM14-NRAS cells were resistant in vitro to either gilteritinib or trametinib alone, but remained sensitive to the two drugs in combination. Both serial single cell genotyping to determine whether NRAS mutations precede gilteritinib therapy and whole exome sequencing of progression samples lacking new mutations by targeted NGS are ongoing and will be updated for presentation.
Conclusion
Ras mutations are the most common mechanism of acquired mutational resistance to gilteritinib and can occur through clonal selection for FLT3-WT or clonal evolution of FLT3-mutated cells. Combining gilteritinib and MEK inhibitors is a promising strategy to prevent or delay clinical resistance.
Session topic: 3. Acute myeloid leukemia - Biology & Translational Research
Keyword(s): Drug resistance, Flt3 inhibitor, Flt3-ITD, Ras
Abstract: S817
Type: Oral Presentation
Presentation during EHA23: On Saturday, June 16, 2018 from 12:30 - 12:45
Location: Room A4
Background
Gilteritinib is a potent, selective FLT3 inhibitor with substantial single-agent clinical activity in relapsed/refractory (R/R) AML patients with FLT3 internal tandem duplication (ITD) and/or FLT3-D835 tyrosine kinase domain mutations (Perl AE, et al Lancet Oncol 2017). However, gilteritinib monotherapy is not curative and, like other FLT3 inhibitors, our data suggest acquired sequence mutations eventually arise and contribute to drug resistance (McMahon CM, et al, ASH 2017 abstract #295). In some cases, FLT3-wild type leukemic populations are selected during gilteritinib therapy and establish clonal dominance at clinical progression; in others, the original FLT3 mutation persists but is accompanied at progression by new mutations, often N-RAS or FLT3-F691L. Higher gilteritinib doses might combat resistance from FLT3-F691L, but combinatorial strategies are likely needed to circumvent other mechanisms of resistance and optimize response quality and duration.
Aims
We sought to describe and clarify mechanisms of secondary resistance to gilteritinib therapy.
Methods
Adults with R/R FLT3-mutated AML who received at least 80 mg/day of gilteritinib on a phase 1/2 study (NCT02014558) at 3 institutions provided leukemia samples at baseline, scheduled study time points, and at study withdrawal; all clinical molecular and cytogenetic data were reviewed. Baseline and progression samples were tested by a 66-gene next generation sequencing (NGS) panel; a subset with treatment-emergent mutations at disease progression were analyzed by a 24-gene single-cell NGS technique (Tapestri). Dual FLT3-ITD+/NRAS+ cell lines (MOLM14-NRAS) were generated through long-term culture of MOLM14 cells in low concentrations of the FLT3 inhibitor quizartinib. Effects of gilteritinib and/or the MEK inhibitor trametinib on the in vitro growth and apoptosis of MOLM14-NRAS cells were studied.
Results
29 subjects (median age 62) have complete, evaluable cytogenetic and/or NGS data; an additional 7 have data currently being analyzed. At study enrollment 26/29 (89.7%) had a FLT3-ITD mutation, including 5 (17.2%) with both FLT3-ITD and FLT3-D835 mutations. Three subjects (10.3%) had a FLT3-D835 mutation only. 23/29 progressed during gilteritinib therapy, 5 were withdrawn for transplant or toxicity, and one remains on gilteritinib in remission. 10/21 subjects (47.6%) with serial cytogenetic data developed new cytogenetic abnormalities during therapy, including a new BCR-ABL1 fusion at progression in 1 subject. Cytogenetic evolution did not always correlate with disease progression. New gene mutations developed on gilteritinib in 12/27 subjects (44.4%) with available NGS data, including NRAS (n=7), KRAS (n=2), FLT3-F691L (n=3), IDH2 (n=1), PTPN11 (n=2), TBL1XR1 (n=1), and CBL (n=1). Single-cell NGS confirmed that treatment-emergent RAS mutations indeed occur in FLT3-mutated cells. MOLM14-NRAS cells were resistant in vitro to either gilteritinib or trametinib alone, but remained sensitive to the two drugs in combination. Both serial single cell genotyping to determine whether NRAS mutations precede gilteritinib therapy and whole exome sequencing of progression samples lacking new mutations by targeted NGS are ongoing and will be updated for presentation.
Conclusion
Ras mutations are the most common mechanism of acquired mutational resistance to gilteritinib and can occur through clonal selection for FLT3-WT or clonal evolution of FLT3-mutated cells. Combining gilteritinib and MEK inhibitors is a promising strategy to prevent or delay clinical resistance.
Session topic: 3. Acute myeloid leukemia - Biology & Translational Research
Keyword(s): Drug resistance, Flt3 inhibitor, Flt3-ITD, Ras