Matrix-regulated integrin αvβ5 maintains α5β1-dependent desmoplastic traits prognostic of neoplastic recurrence

Abstract

Desmoplasia, a fibrotic mass including cancer-associated fibroblasts (CAFs) and self-sustaining extracellular matrix (D-ECM), is a puzzling feature of pancreatic ductal adenocarcinoma (PDACs). Conflicting studies have identified tumor-restricting and tumor-promoting roles of PDAC-associated desmoplasia, suggesting that individual CAF/D-ECM protein constituents have distinguishable tumorigenic and tumor-repressive functions. Using 3D culture of normal pancreatic versus PDAC-associated human fibroblasts, we identified a CAF/D-ECM phenotype that correlates with improved patient outcomes, and that includes CAFs enriched in plasma membrane-localized, active α5β1-integrin. Mechanistically, we established that TGFβ is required for D-ECM production but dispensable for D-ECM-induced naïve fibroblast-to-CAF activation, which depends on αvβ5-integrin redistribution of pFAK-independent active α5β1-integrin to assorted endosomes. Importantly, the development of a simultaneous multi-channel immunofluorescence approach and new algorithms for computational batch-analysis and their application to a human PDAC panel, indicated that stromal localization and levels of active SMAD2/3 and α5β1-integrin distinguish patient-protective from patient-detrimental desmoplasia and foretell tumor recurrences, suggesting a useful new prognostic tool.DOI: http://dx.doi.org/10.7554/eLife.20600.001

Netrin G1 promotes pancreatic tumorigenesis through cancer associated fibroblast driven nutritional support and immunosuppression

“…NetG1 was upregulated in the stromal compartment (pan-cytokeratin -/Epcam -, vimentin + areas) in tumor tissue compared to tumor adjacent normal tissue and normal pancreatic tissue from individuals without cancer (Fig 2A, B). Moreover, stromal tyrosine 397 phosphorylated focal adhesion kinase (pFAK) expression followed a similar pattern of increased expression in tumor tissue compared to normal and tumor adjacent normal tissue, in line with our established pro-tumor CAF signature (Franco-Barraza et al, 2017). We also observed a strong correlation between stromal NetG1 expression and stromal pFAK expression in all tissues combined (R 2 = 0.8046, P< 0.0001) (Fig 2C).…”

“…NetG1 is present on pre-synaptic neurons, and it stabilizes excitatory glutamatergic synapses by interacting with its only known binding partner on post-synaptic cells, Netrin G1 Ligand (NGL-1) (Lin et al, 2003;Song et al, 2013). To assess the clinical relevance of NetG1/NGL-1, we performed Simultaneous Multichannel Immunofluorescence (SMI) (Franco-Barraza et al, 2017) on 6 normal, 4 tumor adjacent and 15 tumor pancreatic tissue samples to discern NetG1 and NGL-1 expression patterns in distinct subsets of cells. NetG1 was upregulated in the stromal compartment (pan-cytokeratin -/Epcam -, vimentin + areas) in tumor tissue compared to tumor adjacent normal tissue and normal pancreatic tissue from individuals without cancer (Fig 2A, B).…”

Targeting the complexity of Src signalling in the tumour microenvironment of pancreatic cancer: from mechanism to therapy

“…In PDAC specifically, overexpression of integrin amb3/ amb6 has been previously shown to associate with poor survival of patients as well as lymph node metastasis [59,72], and recent findings indicate that the stromal localisation and levels of active a5b1-integrin and FAK can identify two readily distinguishable desmoplastic phenotypes in pancreatic cancer. Tumours with high stromal pSMAD2/3 levels were found to be prognostic of poor outcome, whilst increased stromal levels of active a,b-integrin constituted a patient-protective PDAC-associated desmoplastic phenotype [73]. In addition, integrins also play a role in regulating cancer stem cell properties leading to metastasis as well as resistance to tyrosine kinase inhibitors in PDAC [74].…”

Force-exerting lateral protrusions in fibroblastic cell contraction

“…Correlating the alignment of f-actin stress fibers with activation of phosphorylated focal adhesion kinase (pFAK) at adhesion sites on neighboring fibers will provide further evidence on formation of 3D adhesions and cells activating to force generating myofibroblasts. 61 We emphasize that our method of force measurement establishes force vectors originating from adhesion sites and directed along f-actin stress fibers contrasts other methods where the force vectors are determined independently of stress fiber orientation [67][68][69] .…”

“…The parallel fibers used in our method mimic the aligned ECMs produced by cancer associated fibroblasts (CAFs) in vitro and in vivo TACs2/3 fiber configurations known to enable CAF activation 11,61 . The inter-fiber spacing (~10 µm) used in our study mimics those measured by the fibroblastic cell-derived ECMs model and single harmonic generation (SHG) imaging of the patient samples 7,9,11,[61][62][63] . At higher inter-fiber spacing 64,65 , cells attach to single fibers in spindle morphologies with limited instances of twine formation, akin to the behavior reported in elongated spindles in 3D gels 15 .…”