CHAPTER 16. Fibroblastic Cell-derived Extracellular Matrices: A Cell Culturing System to Model Key Aspects of the Tumor Microenvironment

Gardiner, Jaye C.; Raghavan, Kristopher S.; Alexander, Jennifer I.; Franco‐Barraza, Janusz; Cukierman, Edna

doi:10.1039/9781788015998-00305

Cited by 6 publications

(10 citation statements)

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“…These results suggest that eribulin effects on PDAC survival (e.g., Figures 4C and Supplemental 2), are the result of the eribulin-imparted effects on CAFs, as opposed to a direct effect upon PDAC cells. Altogether these observations are akin to reported TGFβ-deficient CAFs, whereby TGFβ signaling was shown to be uniquely necessary to produce a tumor-supportive ECM [15, 22, 33, 40].…”

Section: Resultsmentioning

confidence: 72%

“…In addition to the effects of eribulin on ECM alignment, we questioned whether this drug could hinder known intracellular CAF biomarkers indicative of tumor-supportive CAF function [40]. For this, we quantified the levels of expression of the actin-bundling protein palladin, as well as constitutive pSmad2/3 activity.…”

Section: Resultsmentioning

confidence: 99%

“…Similarly to using TGFR1i or β5 KO CAFs, increased doses of eribulin (e.g., 1 nM and 3 nM) triggered 30% and 50% decrease in palladin expression (0.73 ± 0.13 and 0.51 ± 0.14) respectively, with analogous tendencies for pSmad2/3 (0.67 ± 0.27 and 0.53 ± 0.21), compared to vehicle control (Figures 2C and Supplemental 1C). We also queried the effects of eribulin in the levels of focal adhesion kinase tyrosine-397 phosphorylation (pFAK), as well as intensities of the active conformation of the fibronectin receptor α5β1-integrin, as these two markers are too indicative of tumor-supportive CAF function in vitro (e.g., in the 3D CDM system), and have been associated with worse patient outcomes in vivo [15, 40]. Although levels of pFAK solely showed a modest tendency of 20% reduction in response to 1 nM eribulin (81% ± 25%), a significant decrease of 34% (66% ± 12%) in the activity of this kinase was evident with 3 nM eribulin (Figure 2D).…”

Section: Resultsmentioning

confidence: 99%

“…Note that all tendencies were similar to results obtained with TGFR1i and in β5 KO CAFs, yet paclitaxel did not affect the gauged pFAK or active α5β1-integrin levels. Altogether, these data suggest that eribulin triggers a phenotypic transition from tumor-supportive to tumor-suppressive, which is like known TGFβ inhibition and thus renders CAFs with features akin to “normalized” β5 KO CAFs [15, 40].…”

Section: Resultsmentioning

confidence: 99%

“…The goal of this study was hence to assess the in vitro modulating effects that eribulin imparts on PDAC CAF function. For this, we used our well-characterized in vivo -mimetic desmoplastic system, referred to in this study as the CAF-generated CDM system, which is ideally suited for testing CAF phenotypes and functions [15, 23, 40].…”

Section: Discussionmentioning

confidence: 99%

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Eribulin normalizes pancreatic cancer-associated fibroblasts by simulating TGFβ inhibition

Luong

Cukierman

2022

Preprint

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Less than 11% of pancreatic cancer patients survive 5-years post-diagnosis. The unique biology of pancreatic cancer includes a significant expansion of its desmoplastic tumor microenvironment, wherein cancer-associated fibroblasts (CAFs) and their self-produced extracellular matrix are key components. CAF functions are both tumor-supportive and tumor-suppressive, while normal fibroblastic cells are solely tumor-suppressive. Knowing that CAF-eliminating drugs are ineffective and can accelerate cancer progression, therapies that normalize CAF function are highly pursued. Eribulin is a well-tolerated anti-microtubule drug used to treat a plethora of neoplasias, including advanced/metastatic cancers. Importantly, eribulin can inhibit epithelial to mesenchymal transition via a mechanism akin to blocking pathways induced by transforming growth factor-beta (TGFb). Notably, canonical TGFb signaling also plays a pivotal role in CAF activation, which is necessary for the development and maintenance of desmoplasia. Hence, we hypothesized that eribulin could modulate, and perhaps normalize CAF function. To test this premise, we used a well-established in vivo-mimetic fibroblastic cell-derived extracellular matrix (CDM) system and gauged the effects of eribulin on human pancreatic CAFs and cancer cells. The pathophysiologic cell-culturing system was also used to query eribulin effects on CDM-regulated cancer cell survival and invasive spread. Results demonstrated that intact CDMs modestly restricted eribulin from obstructing cancer cell growth. Nonetheless, eribulin-treated CAFs generated CDMs that limited cancer cell survival under nutritional stress, similar to reported tumor-suppressive CDMs generated by TGFb-deficient CAFs. Data from this study support the central proposed premise and suggest that eribulin could be used as a CAF-normalizing drug.

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Section: Resultsmentioning

confidence: 72%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 3 more Smart Citations

Eribulin normalizes pancreatic cancer-associated fibroblasts by simulating TGFβ inhibition

Luong

Cukierman

2022

Preprint

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Exploring the potential role of palladin in modulating human CAF/ECM functional units

Dolskii,

Alcantara dos Santos,

Andrake

et al. 2024

Cytoskeleton

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Fibroblasts, crucial for maintaining tissue homeostasis, significantly shape the tumor microenvironment (TME). In pancreatic cancer, a highly aggressive malignancy, cancer‐associated fibroblast (CAF)/extracellular matrix (ECM) units dominate the TME, influencing tumor initiation, progression, and treatment responses. Palladin, an actin‐associated protein, is vital for fibroblast structural integrity and activation, playing a key role in CAF/ECM functionality. Palladin interacts with cytoskeletal proteins such as alpha‐actinin (α‐Act) and can therefore regulate other proteins like syndecans, modulating cytoskeletal features, cell adhesion, integrin recycling, and signaling. In this review, we propose that targeting the palladin/α‐Act/syndecan interaction network could modulate CAF/ECM units, potentially shifting the TME from a tumor‐promoting to a tumor‐suppressive state. In silico data and reported studies to suggest that stabilizing palladin‐α‐Act interactions, via excess palladin, influences syndecan functions; potentially modulating integrin endocytosis via syndecan engagement with protein kinase C alpha as opposed to syndecan binding to α‐Act. This mechanism can then affect the distribution of active α5β1‐integrin between the plasma membrane and known intracellular vesicular compartments, thereby influencing the tumor‐suppressive versus tumor‐promoting functions of CAF/ECM units. Understanding these interactions offers likely future therapeutic avenues for stroma normalization in pancreatic and other cancers, aiming to inhibit tumor progression and improve future treatment outcomes.

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Computational and experimental characterization of the novel ECM glycoprotein SNED1 and prediction of its interactome

Vallet

Davis

Barqué

et al. 2021

Biochemical Journal

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The extracellular matrix (ECM) is a complex meshwork of proteins and an essential component of multicellular life. We have recently reported the characterization of a novel ECM protein, SNED1, and showed that it promotes breast cancer metastasis and regulates craniofacial development. However, the mechanisms by which it does so remain unknown. ECM proteins exert their functions by binding to cell surface receptors and interacting with other ECM proteins, actions that we can predict using knowledge of protein’s sequence, structure, and post-translational modifications. Here, we combined in-silico and in-vitro approaches to characterize the physico-chemical properties of SNED1 and infer its putative functions. To do so, we established a mammalian cell system to produce and purify SNED1 and its N-terminal fragment, which contains a NIDO domain and demonstrated experimentally SNED1’s potential to be glycosylated, phosphorylated, and incorporated into an insoluble ECM. We also determined the secondary and tertiary structures of SNED1 and its N-terminal fragment and obtained a model for its NIDO domain. Using computational predictions, we identified 114 proteins as putative SNED1 interactors, including the ECM protein fibronectin. Pathway analysis of the predicted SNED1 interactome further revealed that it may contribute to signaling through cell surface receptors, such as integrins, and participate in the regulation of ECM organization and developmental processes. Last, using fluorescence microscopy, we showed that SNED1 forms microfibrils within the ECM and partially colocalizes with fibronectin. Altogether, we provide a wealth of information on an understudied yet important ECM protein with the potential to decipher its pathophysiological functions.

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CHAPTER 16. Fibroblastic Cell-derived Extracellular Matrices: A Cell Culturing System to Model Key Aspects of the Tumor Microenvironment

Cited by 6 publications

References 0 publications

Eribulin normalizes pancreatic cancer-associated fibroblasts by simulating TGFβ inhibition

Eribulin normalizes pancreatic cancer-associated fibroblasts by simulating TGFβ inhibition

Exploring the potential role of palladin in modulating human CAF/ECM functional units

Computational and experimental characterization of the novel ECM glycoprotein SNED1 and prediction of its interactome

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