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. Author manuscript; available in PMC: 2023 Jul 6.
Published in final edited form as: J Cell Physiol. 2020 Jan 8;235(11):7889–7899. doi: 10.1002/jcp.29443

FIGURE 1.

FIGURE 1

Neratinib exposure reduces the phosphorylation of PAK1, Ezrin, and Merlin, and increases the phosphorylation of MAP4K4. (a) FCC1199 GEMM and PDX human pancreatic cancer cells were transfected with scrambled siRNA molecules or with validated siRNA molecules to knock down the expression of Rubicon. Twenty-four hours after transfection, cells were treated with vehicle control, neratinib (50 nM), sodium valproate (250 μM), or the drugs in combination as indicated for 6 hr. Cells were fixed in place and immunostaining performed to determine the total expression of the indicated proteins and the phosphorylation of these proteins. The data as presented are protein phosphorylation corrected for total protein expression. (n = 3+/−SD) *p < .05 less than vehicle control; #p < .05 greater than vehicle control value. (b) PDX human pancreatic cancer cells were treated with vehicle control or with neratinib (50 nM) for 6 hr. Cells were fixed in place and immunostaining performed to determine the total expression of the indicated proteins and the phosphorylation of these proteins. The data as presented are protein phosphorylation corrected for total protein expression. (n = 3+/−SD) *p < .05 less than vehicle control; #p <.05 greater than vehicle control value. (c) PANC1 and MiaPaca2 cells were transfected with a scrambled siRNA control or with validated siRNA molecules to knock down the expression of Rubicon or Merlin. Twenty-four hours after transfection cells were treated with vehicle control or with (neratinib [50 nM]+sodium valproate [250 μM]) for 24 hr. Cells were isolated, and viability determined via a trypan blue exclusion assay (n = 3+/−SD) #p < .05 greater than the corresponding value in siSCR cells; *p < .05 less than the corresponding value in siSCR cells. (d) PANC1 and MiaPaca2 cells were transfected with a scrambled siRNA control or with validated siRNA molecules to knock down the expression of Rubicon or Merlin. Twenty-four hours after transfection cells were treated with vehicle control or with neratinib (50 nM) for 6 hr. Cells were fixed in place and immunostaining performed to determine the total expression of the indicated proteins and the phosphorylation of these proteins. The data as presented are protein phosphorylation corrected for total protein expression. (n = 3+/−SD) *p < .05 less than vehicle control value; **p < .05 less than siSCR control value; #p < .05 greater than siSCR control value. siSCR, small interfering scrambled; siRNA, small interfering RNA