Figure 7. High SYK activation impairs the targeting of FLT3-ITD-driven AML with small-molecule inhibitors in vitro and in vivo.

(A) Kaplan-Meier curves showing overall survival of mice (n=4) secondary transplanted with double tomato and GFP positive myeloid cells expressing FLT3-ITD in combination with MIT empty, SYK WT, or SYK-TEL (clones from two different donor mice). Statistical significance determined by Log-Rank (Mantel-Cox) test.

(B) H&E staining of the bone marrow and liver of a representative moribund mouse from each indicated group.

(C and D) Growth inhibition of secondary transplantable murine primary and Ba/F3 cells co-expressing FLT3-ITD and indicated SYK constructs and treated with increasing doses of either AC220 (C) or R406 (D). Values are shown relative to day zero (time of seeding), with error bars representing the mean +/− SD.

(E and F) Kaplan-Meier curves showing overall survival of mice (n=6) secondary transplanted with double tomato and GFP positive myeloid cells expressing FLT3-ITD in combination with SYK WT (E), or SYK-TEL (F) and treated with vehicle, R788 (R406 prodrug, Fostamatinib), PRT062607, AC220 or a combination of PRT062607 + AC220. Statistical significance determined by Log-Rank (Mantel-Cox) test. ^#, p < 0.02, *, p < 0.002 and **, p ≤ 0.0008 by comparison with vehicle-treated group.

(G) WBC count in the blood harvested from five mice per group at day 10 post-treatment.

(H) Proportion of double tomato and GFP positive secondary transplanted FLT3-ITD + SYK-TEL cells in spleen of five mice sacrificed at day 15 post-treatment. (G and H) p value calculated using a Mann-Whitney test in comparison with vehicle condition. n.s = non significant (p > 0.05). Error bars represent mean +/− SEM.

(I) Western blot showing the levels of FLT3 and SYK phosphorylation on FLT3-ITD + SYK-TEL cells sorted from spleen of mice treated for three days with indicated compounds.

See also Figure S7