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. 2018 Jul 12;13(7):e0200163. doi: 10.1371/journal.pone.0200163

Fig 5. MAPK and AKT pathway inactivation by cetuximab-KRAS- and cetuximab-PIK3CA-esiRNA treatment.

Fig 5

Tumor lysate samples from xenograft experiments were processed for SDS PAGE and probed in Western blots for expression of KRAS downstream MAPK pathway effector phospho-ERK and PIK3CA downstream phospho-AKT along with their unphosphorylated (total) counterparts in DLD1 (A) and HT29 (B) tumors. In DLD1 tumors (A) treated with C-KRAS-, C-PIK3CA- and C-KRAS/PIK3CA-esiRNA, there was a clear reduction of ERK phosphorylation visible in contrary to a control siRNA treatment (top row) and compared to total ERK levels, whereas BRAF-mutated HT29 tumors (B) treated with C-KRAS-esiRNA did not response in terms of ERK phosphorylation. Here, C-PIK3CA- and C-KRAS/PIK3CA-esiRNA treatment elicited reduced phosphorylation. The same western blot membranes were stripped and probed for phosphorylated AKT (p-AKT, third row from above) and total AKT (fourth row from above). Here, C-KRAS-esiRNA treatment did not change AKT phosphorylation in DLD1 (A) as well as HT29 (B) tumors as compared to control siRNA treatment, but C-PIK3CA-esiRNA treatment and C-KRAS/PIK3CA-esiRNA markedly reduced phosphorylation of AKT indicating deactivation of PI3K signaling pathway. Actin was detected as loading control for total protein.