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Fig. 3
Pharmacological inhibition of candidates from the phosphoproteomic analysis reveals NPM1 as potentially being downstream of neratinib-induced neutrophil apoptosis. Human neutrophils were incubated with inhibitors of SIRT1 or NPM1, alone or in combination with 25 μM of neratinib, for 6 h, after which the levels of apoptosis were assessed by morphology. In both experiments, neratinib alone induced neutrophil apoptosis in comparison to the DMSO control, as expected. The inhibitor of SIRT1 (EX527) did not induce human neutrophil apoptosis (A). The inhibitor of NPM1 (NSC348884) did induce human neutrophil apoptosis (B), and no additional apoptosis was observed when this inhibitor was used in combination with neratinib. Each data point shows data from one healthy donor (n = 3); bars represent mean ± standard deviation. One-way ANOVA with multiple comparisons was used to calculate statistical significance; p-values were indicated.