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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Research Article Oncology

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 2

CCL2 induced by TGF-α is enriched in cocultures of cancer cells and DRG neurons, TPAC neurons, and in patient-derived PDAC samples with neural invasion.

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CCL2 induced by TGF-α is enriched in cocultures of cancer cells and DRG ...
(A) Volcano plot of differentially expressed genes (DEGs) for the comparison of murine cancer cells derived from TPAC versus KPC mice. (B) Bar plot displaying a selected set of over-represented pathways among the DEGs for the comparison groups TPAC versus KPC. (C) Pathway-based interaction networks of DEGs for the comparison groups TPAC versus KPC, where network line thickness indicates the confidence of the interaction. (D) Experimental set-up: DRGs from neonatal mice were cultured for 2 days, and rTGF-α was added at concentrations of 1, 10, 25, 50, and 100 ng/mL for 24 hours. After the treatment, cells were used for RT-qPCR. (E) Graphs representing Ccl2 and Npy mRNA content in DRGs after treatment with rTGF-α. (F) Representative images of pancreatic tumors from TPAC and KPC mice stained with PGP9.5 and CCL2 (both in brown). (G) Plots showing the colorimetric CCL2 content in nerves from TPAC and KPC tumors. (H) Representative images of consecutive sections of PDAC patient samples stained with CCL2 and S100 (both in brown) and counterstained with H&E. (I) Plots showing the colorimetric CCL2 content in the nerves of patient samples measured with the QuPath software. Scale bars: 50 μm. All results in the graphs are shown as mean ± SEM. For statistical analyses, ordinary 1-way ANOVA (E), Kruskal-Wallis test (E), Dunnett’s test for multiple comparisons (E), Mann-Whitney U test (G and I), and Shapiro-Wilk normality test for distribution (all panels) were used.