PI3K regulation of RAC1 is required for KRAS-induced pancreatic tumorigenesis in mice

Chia Yen C. Wu; Eileen S. Carpenter; Kenneth K. Takeuchi; Christopher J. Halbrook; Louise V. Peverley; Harold Bien; Jason C. Hall; Kathleen E. Delgiorno; Debjani Pal; Yan Song; Chanjuan Shi; Richard Z. Lin; Howard C. Crawford

doi:10.1053/j.gastro.2014.08.032

PI3K regulation of RAC1 is required for KRAS-induced pancreatic tumorigenesis in mice

Chia Yen C. Wu, Eileen S. Carpenter, Kenneth K. Takeuchi, Christopher J. Halbrook, Louise V. Peverley, Harold Bien, Jason C. Hall, Kathleen E. Delgiorno, Debjani Pal, Yan Song, Chanjuan Shi, Richard Z. Lin, Howard C. Crawford

Cancer Biology

Research output: Contribution to journal › Article › peer-review

65 Scopus citations

Abstract

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.

Original language	English (US)
Pages (from-to)	1405-1416.e7
Journal	Gastroenterology
Volume	147
Issue number	6
DOIs	https://doi.org/10.1053/j.gastro.2014.08.032
State	Published - Dec 1 2014

Keywords

Cytoskeleton
Oncogene
Pancreatic Cancer
Signal Transduction

ASJC Scopus subject areas

Hepatology
Gastroenterology

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10.1053/j.gastro.2014.08.032

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@article{8d728d6648a1476c8d2fbd1994eac29e,

title = "PI3K regulation of RAC1 is required for KRAS-induced pancreatic tumorigenesis in mice",

abstract = "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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ mice.",

keywords = "Cytoskeleton, Oncogene, Pancreatic Cancer, Signal Transduction",

author = "Wu, {Chia Yen C.} and Carpenter, {Eileen S.} and Takeuchi, {Kenneth K.} and Halbrook, {Christopher J.} and Peverley, {Louise V.} and Harold Bien and Hall, {Jason C.} and Delgiorno, {Kathleen E.} and Debjani Pal and Yan Song and Chanjuan Shi and Lin, {Richard Z.} and Crawford, {Howard C.}",

note = "Publisher Copyright: {\textcopyright} 2014 AGA Institute.",

year = "2014",

month = dec,

day = "1",

doi = "10.1053/j.gastro.2014.08.032",

language = "English (US)",

volume = "147",

pages = "1405--1416.e7",

journal = "Gastroenterology",

issn = "0016-5085",

publisher = "W.B. Saunders Ltd",

number = "6",

}

TY - JOUR

T1 - PI3K regulation of RAC1 is required for KRAS-induced pancreatic tumorigenesis in mice

AU - Wu, Chia Yen C.

AU - Carpenter, Eileen S.

AU - Takeuchi, Kenneth K.

AU - Halbrook, Christopher J.

AU - Peverley, Louise V.

AU - Bien, Harold

AU - Hall, Jason C.

AU - Delgiorno, Kathleen E.

AU - Pal, Debjani

AU - Song, Yan

AU - Shi, Chanjuan

AU - Lin, Richard Z.

AU - Crawford, Howard C.

PY - 2014/12/1

Y1 - 2014/12/1

N2 - 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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ mice.

AB - 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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ 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 KrasG12D/+;Ptf1aCre/+ mice.

KW - Cytoskeleton

KW - Oncogene

KW - Pancreatic Cancer

KW - Signal Transduction

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U2 - 10.1053/j.gastro.2014.08.032

DO - 10.1053/j.gastro.2014.08.032

M3 - Article

C2 - 25311989

AN - SCOPUS:84913549563

SN - 0016-5085

VL - 147

SP - 1405-1416.e7

JO - Gastroenterology

JF - Gastroenterology

IS - 6

ER -

PI3K regulation of RAC1 is required for KRAS-induced pancreatic tumorigenesis in mice

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