Kaempferol potentiates the sensitivity of pancreatic cancer cells to erlotinib via inhibition of the PI3K/AKT signaling pathway and epidermal growth factor receptor

Abstract

Erlotinib (ERL) is an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) of pancreatic cancer (PC). However, the clinical efficacy of ERL is limited due to the activation of alternative pathways that bypass the EGFR signaling. Kaempferol (KAE), a natural flavonoid compound, has been reported to possess potent anti-tumor and anti-inflammatory properties, and in this study, we aimed at identifying the sensitization effect of KAE on ERL monotherapy in PC cells and mouse models. Briefly, the CCK-8, colony formation, and flow cytometry were used to assess the proliferation and apoptosis of two PC cell lines in response to a treatment combination of KAE and ERL. Additionally, the drug-disease targets and related anti-PC mechanisms of KAE and ERL were predicted with a network pharmacology method. The survival outcome for PC patients with EGFR differential expression was evaluated through survival analysis. The molecular docking technique predicted the affinity between KAE and EGFR. Moreover, western blot (WB) and immunohistochemistry (IHC) analyses were applied to verify the expression levels of related proteins. As a result, in vitro results showed that the combination of KAE and ERL significantly inhibited cell proliferation and promoted cell apoptosis compared to that with ERL alone. Network pharmacology results demonstrated that KAE sensitized PC to ERL treatment may likely be related to the PI3K/AKT signaling pathway and EGFR TKI resistance. Survival analysis illustrated that PC patients with high expression of EGFR had a relative lower survival rate. Molecular docking results further suggested that KAE had a high binding affinity of - 8.9 kcal/mol with EGFR. WB results indicated that the combination of KAE and ERL dramatically downregulated the expression levels of p-EGFR, p-AKT, p-ERK1/2, and Bcl-2, and upregulated the expression levels of cleaved caspase-9, cleaved PARP, and Bax. The in vivo results revealed that treatment combination of KAE and ERL further reduced the volume and weight of subcutaneous grafted tumors. IHC results confirmed the WB results. These data imply that KAE may be a valid therapeutic candidate to potentiate PC cell sensitivity to ERL via inhibiting PI3K/AKT and EGFR signaling.

Keywords: Epidermal growth factor receptor; Erlotinib; Kaempferol; PI3K/AKT Signaling pathway; Pancreatic cancer; Tumor sensitivity.