Activation of focal adhesion kinase enhances the adhesion and invasion of pancreatic cancer cells via extracellular signal-regulated kinase-1/2 signaling pathway activation.
BACKGROUND
Interaction with integrin and focal adhesion kinase (FAK) regulates the cancer cell adhesion and invasion into extracellular matrix (ECM). In addition, phosphorylation of FAK correlates with the increase of cell motility and invasion. Adhesion and spreading of cancer cells on a variety of ECM proteins, including collagen type IV (Coll IV), leads to an increase in tyrosine phosphorylation and activation of FAK. In this study, we investigated the mechanism of activation of FAK and its downstream extracellular signal-regulated kinase (ERK)-1/2 signaling following stimulation by interleukin (IL)-1alpha and adhesion to ECM with subsequent enhancement of pancreatic cancer cell adhesion and invasion.


RESULTS
In immunoblotting analysis, all three pancreatic cancer cell lines (AsPC-1, BxPC-3, and Capan-2) expressed the protein of FAK and beta1 integrin. Enhancement of FAK protein association with beta1 integrin when cells were plated on Coll IV was more increased by stimulation with IL-1alpha. Preincubation with anti-beta1 integrin antibody and FAK siRNA transfection inhibited the association of FAK with beta1 integrin of pancreatic cancer cells. FAK phosphorylation was observed by adhesion to Coll IV, furthermore, stronger FAK phosphorylation was observed by stimulation with IL-1alpha of pancreatic cancer cells adhered to Coll IV in time-dependent manner. Genistein, a tyrosine kinase inhibitor, markedly inhibited the FAK phosphorylation. IL-1alpha stimulation and Coll IV adhesion enhanced the activation of Ras, as evidenced by the increased Ras-GTP levels in pancreatic cancer cells. Activation of Ras correlated with the phosphorylation of ERK. While not statistical affecting the apoptosis of pancreatic cancer cells, IL-1alpha-induced adhesion and invasion on Coll IV were inhibited with FAK gene silencing by siRNA, beta1 integrin blocking, and inhibition of FAK phosphorylation. PD98059, a MEK inhibitor, also inhibited IL-1alpha-induced enhancement of adhesion and invasion in pancreatic cancer cells.


CONCLUSIONS
Our results demonstrated that activation of FAK is involved with the aggressive capability in pancreatic cancer through Ras/ERK signaling pathway. Based on our results, we suggest that the modification of IL-1, FAK, and integrins functions might be a novel therapeutic approach to aggressive spread of pancreatic cancer.
Sawai
Hirozumi
H
Department of Gastroenterological Surgery, Nagoya City University Graduate School of Medical Sciences, Nagoya 4678601, Japan. cb8h-swi@asahi-net.or.jp
Okada
Yuji
Y
Funahashi
Hitoshi
H
Matsuo
Yoichi
Y
Takahashi
Hiroki
H
Takeyama
Hiromitsu
H
Manabe
Tadao
T
D017209:Apoptosis; D002448:Cell Adhesion / Q000187:drug effects; D045744:Cell Line, Tumor; D024141:Collagen Type IV / Q000494:pharmacology; D004789:Enzyme Activation; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism; D053582:Interleukin-1alpha / Q000494:pharmacology; D020935:MAP Kinase Signaling System; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009361:Neoplasm Invasiveness; D010190:Pancreatic Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding; D021122:Protein Subunits / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D018631:ras Proteins / Q000378:metabolism
D024141:Collagen Type IV; D019012:Integrin beta1; D053582:Interleukin-1alpha; D021122:Protein Subunits; D034741:RNA, Small Interfering; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D018631:ras Proteins