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R.S. Mali et al.
FLT3-ITD+ model whereas venetoclax prolonged survival of the FLT3-WT model (Figure 2B). Venetoclax combined with quizartinib further improved survival of the FLT3- ITD+ model compared to quizartinib alone, while no com- bination effect was observed in the FLT3-WT model (Figure 2B). Analysis of leukemia cells from vehicle-treated mice showed that FLT3-WT cells had elevated BCL-2 protein expression relative to BCL-XL or MCL-1, which may account for venetoclax sensitivity. Conversely, the FLT3- ITD+ cells showed elevated MCL-1 expression compared to BCL-2 or BCL-XL, consistent with the lack of single agent venetoclax activity (Online Supplementary Figure S3B).
A cohort of drug-treated mice (n=3/group) were assessed at the end of dosing. Venetoclax reduced splenomegaly and leukemic cells in peripheral blood (PB), BM and spleen (Figure 3A-B; Online Supplementary Figure S3C) in the FLT3-WT model. In the FLT3-ITD+ model,
quizartinib treatment significantly reduced splenomegaly and leukemic blasts in the PB, BM and spleen while vene- toclax demonstrated little activity (Figure 3A-B; Online Supplementary Figure S3C). Although quizartinib monotherapy profoundly reduced FLT3-ITD+ PB blasts, significant blasts were still present in the BM (~50%) and spleen (~12%; Figure 3B; Online Supplementary Figure S3C). Notably, the addition of venetoclax to quizartinib reduced leukemic cells in the BM and spleen to less than 1% (Figure 3B; Online Supplementary Figure S3C). Treatment was well tolerated as determined by minimal changes in body weight (Online Supplementary Figure S3D). These results further demonstrate that venetoclax, when com- bined with quizartinib, is more efficacious than either sin- gle agent alone in FLT3-ITD+ primary AML localized within a biologically relevant tumor microenvironment at clinically achievable doses.
A
B
CD
Figure 5. Venetoclax synergistically combined with quizartinib in FLT3-ITD+ cell lines. (A) Cell lines were treated for 48 hours with venetoclax, quizartinib or the com- bination at indicated concentrations. ATP content was determined by CellTiter-Glo and Bliss sums were calculated and plotted for each cell line. Bliss sum of >100 is highly synergistic. (B) Cell lines were treated for 48 hours with quizartinib, venetoclax or the combination as indicated and cell viability was assessed by CellTiter- Glo. Values are normalized to the average of the untreated samples for each cell line. (C) Fms-like tyrosine kinase 3 (FLT3) internal tandem duplication (FLT3-ITD+) cell lines were treated for 48 hours with combinations of quizartinib and venetoclax, AMG 176 or A1331852 as indicated. ATP content was determined by CellTiter- Glo and Bliss sums were calculated and plotted for each cell line. (D) FLT3-ITD+ cell lines were treated for 48 hours with combinations of venetoclax, AMG 176, A1331852 or navitoclax as indicated. ATP content was determined by CellTiter-Glo and Bliss sums were calculated and plotted for each cell line.
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haematologica | 2021; 106(4)