Phenotype screens of murine pancreatic cancer identify a Tgf-α-Ccl2-paxillin axis driving human-like neural invasion
Xiaobo Wang, … , Güralp Onur Ceyhan, Ihsan Ekin Demir
Xiaobo Wang, … , Güralp Onur Ceyhan, Ihsan Ekin Demir
Published August 22, 2023
Citation Information: J Clin Invest. 2023;133(21):e166333. https://doi.org/10.1172/JCI166333.
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Phenotype screens of murine pancreatic cancer identify a Tgf-α-Ccl2-paxillin axis driving human-like neural invasion

Abstract

Solid cancers like pancreatic ductal adenocarcinoma (PDAC), a type of pancreatic cancer, frequently exploit nerves for rapid dissemination. This neural invasion (NI) is an independent prognostic factor in PDAC, but insufficiently modeled in genetically engineered mouse models (GEMM) of PDAC. Here, we systematically screened for human-like NI in Europe’s largest repository of GEMM of PDAC, comprising 295 different genotypes. This phenotype screen uncovered 2 GEMMs of PDAC with human-like NI, which are both characterized by pancreas-specific overexpression of transforming growth factor α (TGF-α) and conditional depletion of p53. Mechanistically, cancer-cell-derived TGF-α upregulated CCL2 secretion from sensory neurons, which induced hyperphosphorylation of the cytoskeletal protein paxillin via CCR4 on cancer cells. This activated the cancer migration machinery and filopodia formation toward neurons. Disrupting CCR4 or paxillin activity limited NI and dampened tumor size and tumor innervation. In human PDAC, phospho-paxillin and TGF-α–expression constituted strong prognostic factors. Therefore, we believe that the TGF-α-CCL2-CCR4-p-paxillin axis is a clinically actionable target for constraining NI and tumor progression in PDAC.

Authors

Xiaobo Wang, Rouzanna Istvanffy, Linhan Ye, Steffen Teller, Melanie Laschinger, Kalliope N. Diakopoulos, Kıvanç Görgülü, Qiaolin Li, Lei Ren, Carsten Jäger, Katja Steiger, Alexander Muckenhuber, Baiba Vilne, Kaan Çifcibaşı, Carmen Mota Reyes, Ümmügülsüm Yurteri, Maximilian Kießler, Ibrahim Halil Gürçınar, Maya Sugden, Saliha Elif Yıldızhan, Osman Uğur Sezerman, Sümeyye Çilingir, Güldal Süyen, Maximilian Reichert, Roland M. Schmid, Stefanie Bärthel, Rupert Oellinger, Achim Krüger, Roland Rad, Dieter Saur, Hana Algül, Helmut Friess, Marina Lesina, Güralp Onur Ceyhan, Ihsan Ekin Demir

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Figure 6

Inhibition of Paxillin-Src-Erk signalosome in vitro and in vivo.

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Inhibition of Paxillin-Src-Erk signalosome in vitro and in vivo. (A) Exp...
(A) Experimental design: SU86.86 cancer cells pretreated with the paxillin phosphorylation inhibitor 6-B345TTQ for 1 hour were used for a migration assay with murine DRG neurons. Cells pretreated with vehicle were used as control. (B) Graphs showing the FMI, speed, and distance of cells migrating to DRGs. (C) Scheme of in vivo treatment with 6-B345TTQ. 5-month-old TPAC mice were treated with 1 mg/kg of the inhibitor daily for 5 days for a total of 4 weeks. The control group was treated with DMSO. (D) Representative images of pancreatic tumors from TPAC mice treated with 6-B345TTQ and the control group, stained with the neural marker PGP9.5 (pink), cancer cell marker CK-19 (brown), p-paxillin (brown), and counterstained with haematoxilin (blue). The tumor injection bed is marked by the yellow border lines. Graphs show PGP9.5 and CK-19 content (E) and p-paxillin (F) as percentage of positively stained cells stained in the total area. (G) Scheme of treatment in mice allografted with TPAC cancer cells: 1 × 106 cultured primary TPAC cancer cells were orthotopically transplanted into the pancreas of 129xC57Bl6 mice. 3 weeks after transplantation, the mice were treated according to the regimen. (H) Representative images of pancreatic tumors of transplanted TPAC mice treated according to the regimen, stained with H&E (blue/pink), p-paxillin (brown) and counterstained with haematoxilin (blue). (I) Diagrams showing the percentage of tumor area out of the total analyzed area of transplanted mice after treatment. (J) Diagrams showing the p-paxillin content as percentage of positively stained cells in relation to the total area. All results in graphs are shown as a mean value ± SEM. For the statistical analyses we used Mann-Whitney U test (E, F, I, and J), t test (B and E), and for the distribution, Shapiro-Wilk normality test (all panels). The P value ˂ 0.05 was considered to have significance. Scale bars: 20 μm.