Zeb1 in Stromal Myofibroblasts Promotes Kras-Driven Development of Pancreatic Cancer.

The transcription factor Zeb1 has been identified as a crucial player in Kras-dependent oncogenesis. In pancreatic ductal adenocarcinoma (PDAC), Zeb1 is highly expressed in myofibroblasts and correlates with poor prognosis. As Kras mutations are key drivers in PDAC, we aimed here to assess the necessity of Zeb1 for Kras-driven PDAC and to define the role of Zeb1-expressing myofibroblasts in PDAC development. Genetically engineered mice with conditional pancreatic KrasG12D and Trp53 mutations (KPC) were crossed with Zeb1 haploinsufficient mice (Z+/-). Extensive PDAC was prominent in all 20-week-old KPC;Z+/+ mice, whereas only low-grade precursor lesions were detected in age-matched KPC;Z+/- littermates, with PDAC developing eventually in KPC;Z+/- aged animals. Zeb1 expression in myofibroblasts occurred early in tumorigenesis and Zeb1 haploinsufficiency retarded native expansion of stromal myofibroblasts during precursor-to-cancer progression. Zeb1 downregulation in mPSC repressed their activated gene profile, impaired their migratory and proliferative activity, and attenuated their tumor-supporting features. Conditioned media from Z+/+ mouse-activated (myofibroblast-like) pancreatic stellate cells (mPSC) boosted Ras activity in pancreatic cancer cells carrying mutant Kras; this effect was not observed when using conditioned media from Z+/- mPSC, revealing a paracrinal cooperative axis between Zeb1-expressing PSC and oncogenic Kras-bearing tumor cells. We conclude that Zeb1-expressing stromal myofibroblasts enable a heterotypic collaboration with the Kras-fated epithelial compartment, thus supporting pancreatic malignancy.Significance: Zeb1 expression in stromal myofibroblasts supports PDAC development via collaboration with the epithelial compartment bearing oncogenic Kras mutations. Cancer Res; 78(10); 2624-37. ©2018 AACR.

Sangrador I ，Molero X ，Campbell F ，Franch-Expósito S ，Rovira-Rigau M ，Samper E ，Domínguez-Fraile M ，Fillat C ，Castells A ，Vaquero EC ... -  《-》

Nicotine promotes initiation and progression of KRAS-induced pancreatic cancer via Gata6-dependent dedifferentiation of acinar cells in mice.

Although smoking is a leading risk factor for pancreatic ductal adenocarcinoma (PDAC), little is known about the mechanisms by which smoking promotes initiation or progression of PDAC. We studied the effects of nicotine administration on pancreatic cancer development in Kras(+/LSLG12Vgeo);Elas-tTA/tetO-Cre (Ela-KRAS) mice, Kras(+/LSLG12D);Trp53+/LSLR172H;Pdx-1-Cre (KPC) mice (which express constitutively active forms of KRAS), and C57/B6 mice. Mice were given nicotine for up to 86 weeks to produce blood levels comparable with those of intermediate smokers. Pancreatic tissues were collected and analyzed by immunohistochemistry and reverse transcriptase polymerase chain reaction; cells were isolated and assayed for colony and sphere formation and gene expression. The effects of nicotine were also evaluated in primary pancreatic acinar cells isolated from wild-type, nAChR7a(-/-), Trp53(-/-), and Gata6(-/-);Trp53(-/-) mice. We also analyzed primary PDAC cells that overexpressed GATA6 from lentiviral expression vectors. Administration of nicotine accelerated transformation of pancreatic cells and tumor formation in Ela-KRAS and KPC mice. Nicotine induced dedifferentiation of acinar cells by activating AKT-ERK-MYC signaling; this led to inhibition of Gata6 promoter activity, loss of GATA6 protein, and subsequent loss of acinar differentiation and hyperactivation of oncogenic KRAS. Nicotine also promoted aggressiveness of established tumors as well as the epithelial-mesenchymal transition, increasing numbers of circulating cancer cells and their dissemination to the liver, compared with mice not exposed to nicotine. Nicotine induced pancreatic cells to acquire gene expression patterns and functional characteristics of cancer stem cells. These effects were markedly attenuated in K-Ras(+/LSL-G12D);Trp53(+/LSLR172H);Pdx-1-Cre mice given metformin. Metformin prevented nicotine-induced pancreatic carcinogenesis and tumor growth by up-regulating GATA6 and promoting differentiation toward an acinar cell program. In mice, nicotine promotes pancreatic carcinogenesis and tumor development via down-regulation of Gata6 to induce acinar cell dedifferentiation.

Hermann PC ，Sancho P ，Cañamero M ，Martinelli P ，Madriles F ，Michl P ，Gress T ，de Pascual R ，Gandia L ，Guerra C ，Barbacid M ，Wagner M ，Vieira CR ，Aicher A ，Real FX ，Sainz B Jr ，Heeschen C ... -  《-》

A genetically engineered mouse model developing rapid progressive pancreatic ductal adenocarcinoma.

The premalignant lesions of pancreatic cancer, pancreatic intraepithelial neoplasia (PanIN), have a high frequency of mutations in Kirsten rat sarcoma viral oncogene homologue (KRAS), and genetic alterations in the retinoblastoma (Rb)-E2 factor (E2F) and transformed 3T3 cell double minute 2 (MDM2)-p53 pathways accelerate development of pancreatic ductal adenocarcinoma. The viral oncoprotein SV40 large T antigen (TAg) can inhibit the effects of the Rb family of molecules and of p53 on these pathways, and targeted expression of TAg in mouse pancreas is associated with the development of endocrine or acinar cell tumours. In this study, to determine whether the viral oncoprotein promotes pancreatic duct carcinogenesis initiated by oncogenic KRAS, we generated mice expressing temperature-sensitive SV40 large T antigen (tsTAg) on pancreatic epithelial cells in the presence or absence of Kras(G12D) . Mice with pancreas-specific tsTAg expression developed acinar cell dysplasia by 22 weeks without PanIN formation, while mice expressing both tsTAg and Kras(G12D) developed highly aggressive adenocarcinoma with a ductal cell phenotype within a short period, and died within 3 weeks. The tumours resembled human pancreatic ductal adenocarcinoma (PDAC) at the histological level, and oncogenic Kras and tsTAg synergistically activated E2f and Sre transcription in established PDAC cell lines. These results suggest that tsTAg synergistically promotes Kras(G12D) -associated PDAC formation, and our study identifies a new mouse model of PDAC that may allow a better understanding of the mechanism of carcinogenesis in pancreatic carcinoma, which shows a catastrophic clinical course.

Yamaguchi T ，Ikehara S ，Nakanishi H ，Ikehara Y ... -  《-》

Cell of origin affects tumour development and phenotype in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive tumour thought to arise from ductal cells via pancreatic intraepithelial neoplasia (PanIN) precursor lesions. Modelling of different genetic events in mice suggests both ductal and acinar cells can give rise to PDAC. However, the impact of cellular context alone on tumour development and phenotype is unknown. We examined the contribution of cellular origin to PDAC development by inducing PDAC-associated mutations, KrasG12D expression and Trp53 loss, specifically in ductal cells (Sox9CreER;KrasLSL-G12D;Trp53flox/flox ('Duct:KPcKO ')) or acinar cells (Ptf1aCreER;KrasLSL-G12D;Trp53flox/flox ('Acinar:KPcKO ')) in mice. We then performed a thorough analysis of the resulting histopathological changes. Both mouse models developed PDAC, but Duct:KPcKO mice developed PDAC earlier than Acinar:KPcKO mice. Tumour development was more rapid and associated with high-grade murine PanIN (mPanIN) lesions in Duct:KPcKO mice. In contrast, Acinar:KPcKO mice exhibited widespread metaplasia and low-grade as well as high-grade mPanINs with delayed progression to PDAC. Acinar-cell-derived tumours also had a higher prevalence of mucinous glandular features reminiscent of early mPanIN lesions. These findings indicate that ductal cells are primed to form carcinoma in situ that become invasive PDAC in the presence of oncogenic Kras and Trp53 deletion, while acinar cells with the same mutations appear to require a prolonged period of transition or reprogramming to initiate PDAC. Our findings illustrate that PDAC can develop in multiple ways and the cellular context in which mutations are acquired has significant impact on precursor lesion initiation, disease progression and tumour phenotype.

Lee AYL ，Dubois CL ，Sarai K ，Zarei S ，Schaeffer DF ，Sander M ，Kopp JL ... -  《-》

Oncogenic KRAS Reduces Expression of FGF21 in Acinar Cells to Promote Pancreatic Tumorigenesis in Mice on a High-Fat Diet.

Obesity is a risk factor for pancreatic cancer. In mice, a high-fat diet (HFD) and expression of oncogenic KRAS lead to development of invasive pancreatic ductal adenocarcinoma (PDAC) by unknown mechanisms. We investigated how oncogenic KRAS regulates the expression of fibroblast growth factor 21, FGF21, a metabolic regulator that prevents obesity, and the effects of recombinant human FGF21 (rhFGF21) on pancreatic tumorigenesis. We performed immunohistochemical analyses of FGF21 levels in human pancreatic tissue arrays, comprising 59 PDAC specimens and 45 nontumor tissues. We also studied mice with tamoxifen-inducible expression of oncogenic KRAS in acinar cells (KrasG12D/+ mice) and fElasCreERT mice (controls). KrasG12D/+ mice were placed on an HFD or regular chow diet (control) and given injections of rhFGF21 or vehicle; pancreata were collected and analyzed by histology, immunoblots, quantitative polymerase chain reaction, and immunohistochemistry. We measured markers of inflammation in the pancreas, liver, and adipose tissue. Activity of RAS was measured based on the amount of bound guanosine triphosphate. Pancreatic tissues of mice expressed high levels of FGF21 compared with liver tissues. FGF21 and its receptor proteins were expressed by acinar cells. Acinar cells that expressed KrasG12D/+ had significantly lower expression of Fgf21 messenger RNA compared with acinar cells from control mice, partly due to down-regulation of PPARG expression-a transcription factor that activates Fgf21 transcription. Pancreata from KrasG12D/+ mice on a control diet and given injections of rhFGF21 had reduced pancreatic inflammation, infiltration by immune cells, and acinar-to-ductal metaplasia compared with mice given injections of vehicle. HFD-fed KrasG12D/+ mice given injections of vehicle accumulated abdominal fat, developed extensive inflammation, pancreatic cysts, and high-grade pancreatic intraepithelial neoplasias (PanINs); half the mice developed PDAC with liver metastases. HFD-fed KrasG12D/+ mice given injections of rhFGF21 had reduced accumulation of abdominal fat and pancreatic triglycerides, fewer pancreatic cysts, reduced systemic and pancreatic markers of inflammation, fewer PanINs, and longer survival-only approximately 12% of the mice developed PDACs, and none of the mice had metastases. Pancreata from HFD-fed KrasG12D/+ mice given injections of rhFGF21 had lower levels of active RAS than from mice given vehicle. Normal acinar cells from mice and humans express high levels of FGF21. In mice, acinar expression of oncogenic KRAS significantly reduces FGF21 expression. When these mice are placed on an HFD, they develop extensive inflammation, pancreatic cysts, PanINs, and PDACs, which are reduced by injection of FGF21. FGF21 also reduces the guanosine triphosphate binding capacity of RAS. FGF21 might be used in the prevention or treatment of pancreatic cancer.

Luo Y ，Yang Y ，Liu M ，Wang D ，Wang F ，Bi Y ，Ji J ，Li S ，Liu Y ，Chen R ，Huang H ，Wang X ，Swidnicka-Siergiejko AK ，Janowitz T ，Beyaz S ，Wang G ，Xu S ，Bialkowska AB ，Luo CK ，Pin CL ，Liang G ，Lu X ，Wu M ，Shroyer KR ，Wolff RA ，Plunkett W ，Ji B ，Li Z ，Li E ，Li X ，Yang VW ，Logsdon CD ，Abbruzzese JL ，Lu W ... -  《-》