A phosphoproteomic signature of vascular toxicity predicts that bosutinib is nontoxic to human ECs in vitro. Phosphoproteomic profiling was performed on HUVECs treated with each TKI for 3 hours. (A) Marker selection was used to identify the top and bottom 10 peptides that distinguish the toxic (dasatinib, ponatinib, and nilotinib) from the nontoxic (imatinib) TKIs as ranked by signal to noise. Eleven peptides were chosen as the toxicity signature based on P < .05 and q < .25 (bold). (B) The similarity of each drug to one another based on the 11-phosphopeptide signature was calculated using averaged Pearson correlation coefficients. Bosutinib is more similar to the nontoxic TKI, imatinib. The diagonal (each drug vs itself) may be interpreted as replicate reproducibility. (C-D) Leukocyte adhesion (C) was quantified from representative images (D) of adherent fluorescently labeled U937 leukocytes. (E-F) Wound healing (E) was determined by quantifying the number of HUVECs that migrated into the scratch wound from representative images (F). (G) HUVEC survival was quantified by the Cell TiterGlo Luminescent assay. Data in panels C-F are presented as mean fold change ± standard error of the mean relative to vehicle-treated control. Significance was determined by 1-way analysis of variance and Dunnett post-test. *P < .05; ***P < .001 vs vehicle. ##P < .01; ###P < .001 vs dasatinib. The number of experiments per condition is displayed within each bar. (D,F) Original magnification ×4. DMSO, dimethyl sulfoxide.