PI3K Regulation of RAC1 Is Required for KRAS-Induced Pancreatic Tumorigenesis in Mice

• Published in: Gastroenterology (New York, N.Y. 1943) Vol. 147; no. 6; pp. 1405 - 1416.e7
• Main Authors: Wu, Chia-Yen C, Carpenter, Eileen S, Takeuchi, Kenneth K, Halbrook, Christopher J, Peverley, Louise V, Bien, Harold, Hall, Jason C, DelGiorno, Kathleen E, Pal, Debjani, Song, Yan, Shi, Chanjuan, Lin, Richard Z, Crawford, Howard C
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.12.2014
• Subjects: Acinar Cells - cytology; Acinar Cells - metabolism; Adenocarcinoma - genetics; Adenocarcinoma - metabolism; Animals; Carcinogenesis - genetics; Carcinogenesis - metabolism; Carcinoma, Pancreatic Ductal - genetics; Carcinoma, Pancreatic Ductal - metabolism; Cell Transformation, Neoplastic - genetics; Cell Transformation, Neoplastic - metabolism; Class I Phosphatidylinositol 3-Kinases; Cytoskeleton; Cytoskeleton - metabolism; Female; Gastroenterology and Hepatology; Humans; Male; Mice, Mutant Strains; Neuropeptides - genetics; Neuropeptides - metabolism; Oncogene; Pancreatic Cancer; Phosphatidylinositol 3-Kinases - genetics; Phosphatidylinositol 3-Kinases - metabolism; Primary Cell Culture; Proto-Oncogene Proteins p21(ras) - genetics; Proto-Oncogene Proteins p21(ras) - metabolism; rac1 GTP-Binding Protein - genetics; rac1 GTP-Binding Protein - metabolism; Signal Transduction; Signal Transduction - physiology; Transcriptome; mPanIN; PIP 3; IHC; pAKT; ADM; mRNA; phosphate-buffered saline; PDA; EGFR; pancreatic ductal adenocarcinoma; GFP; PBD; messenger RNA; PI3K; glyceraldehyde-3-phosphate dehydrogenase; immunohistochemistry; PDK1; PBS; phospho-AKT; Signal Transduction; pleckstrin homology; Oncogene; acinar-to-ductal metaplasia; epidermal growth factor receptor; qRT-PCR; phosphatidylinositol-3,4,5-trisphosphate; DAPI; 4′,6-diamidino-2-phenylindole; GEF; Pancreatic Cancer; green fluorescent protein; SDS-PAGE; guanine nucleotide exchange factor; p21-binding domain; phosphoinositide-dependent kinase 1; PanIN; quantitative reverse-transcription polymerase chain reaction; PH; Cytoskeleton; pancreatic intraepithelial neoplasia; sodium dodecyl sulfate/polyacrylamide gel electrophoresis; GAPDH; murine pancreatic intraepithelial neoplasia; phosphoinositide 3-kinase; PIP3
• ISSN: 0016-5085; 1528-0012
• DOI: 10.1053/j.gastro.2014.08.032

Background & Aims New drug targets are urgently needed for the treatment of patients with pancreatic ductal adenocarcinoma (PDA). Nearly all PDAs contain oncogenic mutations in the KRAS gene. Pharmacological inhibition of KRAS has been unsuccessful, leading to a focus on downstream effectors that are more easily targeted with small molecule inhibitors. We investigated the contributions of phosphoinositide 3-kinase (PI3K) to KRAS-initiated tumorigenesis. Methods Tumorigenesis was measured in the Kras G12D/+ ;Ptf1a Cre/+ mouse model of PDA; these mice were crossed with mice with pancreas-specific disruption of genes encoding PI3K p110α ( Pik3ca ), p110β ( Pik3cb ), or RAC1 ( Rac1 ). Pancreatitis was induced with 5 daily intraperitoneal injections of cerulein. Pancreata and primary acinar cells were isolated; acinar cells were incubated with an inhibitor of p110α (PIK75) followed by a broad-spectrum PI3K inhibitor (GDC0941). PDA cell lines (NB490 and MiaPaCa2) were incubated with PIK75 followed by GDC0941. Tissues and cells were analyzed by histology, immunohistochemistry, quantitative reverse-transcription polymerase chain reaction, and immunofluorescence analyses for factors involved in the PI3K signaling pathway. We also examined human pancreas tissue microarrays for levels of p110α and other PI3K pathway components. Results Pancreas-specific disruption of Pik3ca or Rac1 , but not Pik3cb , prevented the development of pancreatic tumors in Kras G12D/+ ;Ptf1a Cre/+ mice. Loss of transformation was independent of AKT regulation. Preneoplastic ductal metaplasia developed in mice lacking pancreatic p110α but regressed. Levels of activated and total RAC1 were higher in pancreatic tissues from Kras G12D/+ ;Ptf1a Cre/+ mice compared with controls. Loss of p110α reduced RAC1 activity and expression in these tissues. p110α was required for the up-regulation and activity of RAC guanine exchange factors during tumorigenesis. Levels of p110α and RAC1 were increased in human pancreatic intraepithelial neoplasias and PDAs compared with healthy pancreata. Conclusions KRAS signaling, via p110α to activate RAC1, is required for transformation in Kras G12D/+ ;Ptf1a Cre/+ mice.