Pancreatic ductal adenocarcinomas (PDAs) are characterized by a robust fibroinflammatory response. We show here that this desmoplastic reaction generates inordinately high interstitial fluid ...pressures (IFPs), exceeding those previously measured or theorized for solid tumors, and induces vascular collapse, while presenting substantial barriers to perfusion, diffusion, and convection of small molecule therapeutics. We identify hyaluronan, or hyaluronic acid (HA), as the primary matrix determinant of these barriers and show that systemic administration of an enzymatic agent can ablate stromal HA from autochthonous murine PDA, normalize IFP, and re-expand the microvasculature. In combination with the standard chemotherapeutic, gemcitabine, the treatment permanently remodels the tumor microenvironment and consistently achieves objective tumor responses, resulting in a near doubling of overall survival.
► Extremely high interstitial fluid pressures (IFPs) induce vascular collapse in PDA ► Hyaluronan (HA) is the major determinant of elevated IFP in PDA ► Systemic infusion of an enzyme, PEGPH20, ablates HA from PDA and normalizes IFP ► PEGPH20 + gemcitabine therapy permanently remodels the stroma and increases survival
Organized extracellular matrix (ECM), in the form of aligned architectures, is a critical mediator of directed cancer cell migration by contact guidance, leading to metastasis in solid tumors. ...Current models suggest anisotropic force generation through the engagement of key adhesion and cytoskeletal complexes drives contact-guided migration. Likewise, disrupting the balance between cell–cell and cell–ECM forces, driven by ECM engagement for cells at the tumor–stromal interface, initiates and drives local invasion. Furthermore, processes such as traction forces exerted by cancer and stromal cells, spontaneous reorientation of matrix-producing fibroblasts, and direct binding of ECM modifying proteins lead to the emergence of collagen alignment in tumors. Thus, as we obtain a deeper understanding of the origins of ECM alignment and the mechanisms by which it is maintained to direct invasion, we are poised to use the new paradigm of stroma-targeted therapies to disrupt this vital axis of disease progression in solid tumors.
The notion that cell shape and spreading can regulate cell proliferation has evolved over several years, but only recently has this been linked to forces from within and upon the cell. This emerging ...area of mechanical signaling is proving to be wide-spread and important for all cell types. The microenvironment that surrounds cells provides a complex spectrum of different, simultaneously active, biochemical, structural and mechanical stimuli. In this milieu, cells probe the stiffness of their microenvironment by pulling on the extracellular matrix (ECM) and/or adjacent cells. This process is dependent on transcellular cell-ECM or cell-cell adhesions, as well as cell contractility mediated by Rho GTPases, to provide a functional linkage through which forces are transmitted through the cytoskeleton by intracellular force-generating proteins. This Commentary covers recent advances in the underlying mechanisms that control cell proliferation by mechanical signaling, with an emphasis on the role of 3D microenvironments and in vivo extracellular matrices. Moreover, as there is much recent interest in the tumor-stromal interaction, we will pay particular attention to exciting new data describing the role of mechanical signaling in the progression of breast cancer.
Pancreatic ductal adenocarcinoma (PDA) remains one of the deadliest forms of cancer, in part, because it is largely refractory to current therapies. The failure of most standard therapies in PDA, as ...well as promising immune therapies, may be largely ascribed to highly unique and protective stromal microenvironments that present significant biophysical barriers to effective drug delivery, that are immunosuppressive, and that can limit the distribution and function of antitumor immune cells. Here, we utilized stromal reengineering to disrupt these barriers and move the stroma toward normalization using a potent antifibrotic agent, halofuginone. In an autochthonous genetically engineered mouse model of PDA, halofuginone disrupted physical barriers to effective drug distribution by decreasing fibroblast activation and reducing key extracellular matrix elements that drive stromal resistance. Concomitantly, halofuginone treatment altered the immune landscape in PDA, with greater immune infiltrate into regions of low hylauronan, which resulted in increased number and distribution of both classically activated inflammatory macrophages and cytotoxic T cells. In concert with a direct effect on carcinoma cells, this led to widespread intratumoral necrosis and reduced tumor volume. These data point to the multifunctional and critical role of the stroma in tumor protection and survival and demonstrate how compromising tumor integrity to move toward a more normal physiologic state through stroma-targeting therapy will likely be an instrumental component in treating PDA. SIGNIFICANCE: This work demonstrates how focused stromal re-engineering approaches to move toward normalization of the stroma disrupt physical barriers to effective drug delivery and promote antitumor immunity.
.
The architecture of the extracellular matrix (ECM) directs cell behavior by providing spatial and mechanical cues to which cells respond. In addition to soluble chemical factors, physical ...interactions between the cell and ECM regulate primary cell processes, including differentiation, migration, and proliferation. Advances in microtechnology and, more recently, nanotechnology provide a powerful means to study the influence of the ECM on cell behavior. By recapitulating local architectures that cells encounter in vivo, we can elucidate and dissect the fundamental signal transduction pathways that control cell behavior in critical developmental, physiological, and pathological processes.
Bringing order to the matrix Provenzano, Paolo P
Nature materials,
02/2020, Letnik:
19, Številka:
2
Journal Article
Recenzirano
Odprti dostop
Aligned anisotropic organization of the extracellular matrix by fibroblasts has now been shown to depend on cell reorientation following collision, with the cell collision guidance dependent on the ...transcription factor, TFAP2C.
Directed cell migration by contact guidance in aligned collagenous extracellular matrix (ECM) is a critical enabler of breast cancer dissemination. The mechanisms of this process are poorly ...understood, particularly in 3D, in part because of the lack of efficient methods to generate aligned collagen matrices. To address this technological gap, we propose a simple method to align collagen gels using guided cellular compaction. Our method yields highly aligned, acellular collagen constructs with predictable microstructural features, thus providing a controlled microenvironment for in vitro experiments. Quantifying cell behavior in these anisotropic constructs, we find that breast carcinoma cells are acutely sensitive to the direction and extent of collagen alignment. Further, live cell imaging and analysis of 3D cell migration reveals that alignment of collagen does not alter the total motility of breast cancer cells, but simply redirects their migration to produce largely one-dimensional movement. However, a profoundly enhanced motility in aligned collagen matrices is observed for the subpopulation of carcinoma cells with high tumor initiating and metastatic capacity, termed cancer stem cells (CSCs). Analysis of the biophysical determinants of cell migration show that nuclear deformation is not a critical factor associated with the observed increases in motility for CSCs. Rather, smaller cell size, a high degree of phenotypic plasticity, and increased protrusive activity emerge as vital facilitators of rapid, contact-guided migration of CSCs in aligned 3D collagen matrices.
Directed migration by contact guidance is a poorly understood yet vital phenomenon, particularly for carcinoma cell invasion on aligned collagen fibres. We demonstrate that for single cells, aligned ...architectures providing contact guidance cues induce constrained focal adhesion maturation and associated F-actin alignment, consequently orchestrating anisotropic traction stresses that drive cell orientation and directional migration. Consistent with this understanding, relaxing spatial constraints to adhesion maturation either through reduction in substrate alignment density or reduction in adhesion size diminishes the contact guidance response. While such interactions allow single mesenchymal-like cells to spontaneously 'sense' and follow topographic alignment, intercellular interactions within epithelial clusters temper anisotropic cell-substratum forces, resulting in substantially lower directional response. Overall, these results point to the control of contact guidance by a balance of cell-substratum and cell-cell interactions, modulated by cell phenotype-specific cytoskeletal arrangements. Thus, our findings elucidate how phenotypically diverse cells perceive ECM alignment at the molecular level.
Stromal-epithelial interactions are of particular significance in breast tissue as misregulation of these interactions can promote tumorigenesis and invasion. Moreover, collagen-dense breast tissue ...increases the risk of breast carcinoma, although the relationship between collagen density and tumorigenesis is not well understood. As little is known about epithelial-stromal interactions in vivo, it is necessary to visualize the stroma surrounding normal epithelium and mammary tumors in intact tissues to better understand how matrix organization, density, and composition affect tumor formation and progression.
Epithelial-stromal interactions in normal mammary glands, mammary tumors, and tumor explants in three-dimensional culture were studied with histology, electron microscopy, and nonlinear optical imaging methodologies. Imaging of the tumor-stromal interface in live tumor tissue ex vivo was performed with multiphoton laser-scanning microscopy (MPLSM) to generate multiphoton excitation (MPE) of endogenous fluorophores and second harmonic generation (SHG) to image stromal collagen.
We used both laser-scanning multiphoton and second harmonic generation microscopy to determine the organization of specific collagen structures around ducts and tumors in intact, unfixed and unsectioned mammary glands. Local alterations in collagen density were clearly seen, allowing us to obtain three-dimensional information regarding the organization of the mammary stroma, such as radiating collagen fibers that could not have been obtained using classical histological techniques. Moreover, we observed and defined three tumor-associated collagen signatures (TACS) that provide novel markers to locate and characterize tumors. In particular, local cell invasion was found predominantly to be oriented along certain aligned collagen fibers, suggesting that radial alignment of collagen fibers relative to tumors facilitates invasion. Consistent with this observation, primary tumor explants cultured in a randomly organized collagen matrix realigned the collagen fibers, allowing individual tumor cells to migrate out along radially aligned fibers.
The presentation of these tumor-associated collagen signatures allowed us to identify pre-palpable tumors and see cells at the tumor-stromal boundary invading into the stroma along radially aligned collagen fibers. As such, TACS should provide indications that a tumor is, or could become, invasive, and may serve as part of a strategy to help identify and characterize breast tumors in animal and human tissues.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Elevated interstitial fluid pressure can present a substantial barrier to drug delivery in solid tumors. This is particularly true of pancreatic ductal adenocarcinoma, a highly lethal disease ...characterized by a robust fibroinflammatory response, widespread vascular collapse, and hypoperfusion that together serve as primary mechanisms of treatment resistance. Free-fluid pressures, however, are relatively low in pancreatic ductal adenocarcinoma and cannot account for the vascular collapse. Indeed, we have shown that the overexpression and deposition in the interstitium of high-molecular-weight hyaluronan (HA) is principally responsible for generating pressures that can reach 100 mmHg through the creation of a large gel-fluid phase. By interrogating a variety of tissues, tumor types, and experimental model systems, we show that an HA-dependent fluid phase contributes substantially to pressures in many solid tumors and has been largely unappreciated heretofore. We investigated the relative contributions of both freely mobile fluid and gel fluid to interstitial fluid pressure by performing simultaneous, real-time fluid-pressure measurements with both the classical wick-in-needle method (to estimate free-fluid pressure) and a piezoelectric pressure catheter transducer (which is capable of capturing pressures associated with either phase). We demonstrate further that systemic treatment with pegylated recombinant hyaluronidase (PEGPH20) depletes interstitial HA and eliminates the gel-fluid phase. This significantly reduces interstitial pressures and leaves primarily free fluid behind, relieving the barrier to drug delivery. These findings argue that quantifying the contributions of free- and gel-fluid phases to hydraulically transmitted pressures in a given cancer will be essential to designing the most appropriate and effective strategies to overcome this important and frequently underestimated resistance mechanism.
PDF
