ERC Advanced Grant 741350

What Holds Cancer Cells Back?

 

in context of H2020

 

Since decades the bulk of cancer research focusses on the genetic and molecular level. To complement this knowledge, I will focus on the collective behaviour of cancer cells in cell clusters and in the extracellular matrix (ECM). Conventional cancer research tackles issues like genetic changes, signalling pathways or intracellular mechanisms. In contrast, I want to answer the fundamental, mechanistic question: When is a cancer cell jammed or when can it overcome the yield stress to actively „flow“ in a dense microenvironment (ME)? Within the concept of Physics of Cancer, I have brought forward the idea that changes in a cancer cell’s material properties determine its metastatic potential. To build upon this I propose the next breakthrough by determining a predictive phase diagram for unjamming transitions of cancer cells. Cancer cell jamming is quantified by cell speed as a measure of the motile forces and by cellular shape to account for the interplay between cell contractility and adhesion. Our self-propelled Voronoi model (SPV) will conclusively explain whether a cell is jammed by its neighbours or the ECM, thereby overcoming the limitations of existing theories which only apply to specific environments. Building on my leadership in cell biomechanics and the exclusive clinical access to two types of carcinomas (mamma, cervix), I will introduce highly innovative bionic modulators of intracellular mechanics and develop live cancer cell tracking in biopsies as a ground-breaking alternative to vital imaging. While these approaches are perfect to prove that unjamming transitions are key to tumour progression, I will investigate to what extent fluid, i.e. unjammed, tissue behaviour can be detected by magnetic resonance imaging elastography (MRE) as an individual predictive marker for metastasis. Moreover, the results may guide surgeons when considering the local spreading of cancer and thus greatly empower surgery in tumour therapies.

 

Events

Physics of Cancer

The aim of this symposium is by bringing together exceptional researchers in the areas of quantitative cell biology, physical mechanisms of pathology, cancer biology, molecular design, diagnostic systems and beyond to create a forum for the exchange of new ideas and formulation of new solutions.

2021

2020

2019

2018

2017


Publications

T Händler, C Tutmarc, JS Freitag, DM Smith, J Schnauß:

Constraint Release for Reptating Filaments in Semiflexible Networks Depends on Background Fluctuations Abstract:Entangled semiflexible polymer networks are usually described by the tube model, although this concept has not been able to explain all experimental observations. One of its major shortcomings is neglecting the thermal fluctuations of the polymers surrounding the examined test filament, such that disentanglement effects are not captured. In this study, we present experimental evidence that correlated constraint release which has been predicted theoretically occurs in entangled, but not in crosslinked semiflexible polymer networks. By tracking single semiflexible DNA nanotubes embedded both in entangled and crosslinked F-actin networks, we observed different reptation dynamics in both systems, emphasizing the need for a revision of the classical tube theory for entangled polymer solutions.

Polymers 14(4), 707 (2022)   DOI: 10.3390/polym14040707  

 

D Tschodu, B Ulm, K Bendrat, J Lippoldt, P Gottheil, JA Käs, A Niendorf:

Comparative analysis of molecular signatures reveals a hybrid approach in breast cancer: Combining the Nottingham Prognostic Index with gene expressions into a hybrid signature Abstract:The diagnosis of breast cancer-including determination of prognosis and prediction-has been traditionally based on clinical and pathological characteristics such as tumor size, nodal status, and tumor grade. The decision-making process has been expanded by the recent introduction of molecular signatures. These signatures, however, have not reached the highest levels of evidence thus far. Yet they have been brought to clinical practice based on statistical significance in prospective as well as retrospective studies. Intriguingly, it has also been reported that most random sets of genes are significantly associated with disease outcome. These facts raise two highly relevant questions: What information gain do these signatures procure? How can one find a signature that is substantially better than a random set of genes? Our study addresses these questions. To address the latter question, we present a hybrid signature that joins the traditional approach with the molecular one by combining the Nottingham Prognostic Index with gene expressions in a data-driven fashion. To address the issue of information gain, we perform careful statistical analysis and comparison of the hybrid signature, gene expression lists of two commercially available tests as well as signatures selected at random, and introduce the Signature Skill Score-a simple measure to assess improvement on random signatures. Despite being based on in silico data, our research is designed to be useful for the decision-making process of oncologists and strongly supports association of random signatures with outcome. Although our study shows that none of these signatures can be considered as the main candidate for providing prognostic information, it also demonstrates that both the hybrid signature and the gene expression list of the OncotypeDx signature identify patients who may not require adjuvant chemotherapy. More importantly, we show that combining signatures substantially improves the identification of patients who do not need adjuvant chemotherapy.

PLoS ONE 17(2): e0261035 (2022)   DOI: 10.1371/journal.pone.0261035  

 

I Elbalasy, N Wilharm, E Herchenhahn, R Konieczny, SG Mayr, J Schnauß:

From Strain Stiffening to Softening-Rheological Characterization of Keratins 8 and 18 Networks Crosslinked via Electron Irradiation Abstract:Networks of crosslinked keratin filaments are abundant in epithelial cells and tissues, providing resilience against mechanical forces and ensuring cellular integrity. Although studies of in vitro models of reconstituted keratin networks have revealed important mechanical aspects, the mechanical properties of crosslinked keratin structures remain poorly understood. Here, we exploited the power of electron beam irradiation (EBI) to crosslink in vitro networks of soft epithelial keratins 8 and 18 (k8-k18) filaments with different irradiation doses (30 kGy, 50 kGy, 80 kGy, 100 kGy, and 150 kGy). We combined bulk shear rheology with confocal microscopy to investigate the impact of crosslinking on the mechanical and structural properties of the resultant keratin gels. We found that irradiated keratin gels display higher linear elastic modulus than the unirradiated, entangled networks at all doses tested. However, at the high doses (80 kGy, 100 kGy, and 150 kGy), we observed a remarkable drop in the elastic modulus compared to 50 kGy. Intriguingly, the irradiation drastically changed the behavior for large, nonlinear deformations. While untreated keratin networks displayed a strong strain stiffening, increasing irradiation doses shifted the system to a strain softening behavior. In agreement with the rheological behavior in the linear regime, the confocal microscopy images revealed fully isotropic networks with high percolation in 30 kGy and 50 kGy-treated keratin samples, while irradiation with 100 kGy induced the formation of thick bundles and clusters. Our results demonstrate the impact of permanent crosslinking on k8-k18 mechanics and provide new insights into the potential contribution of intracellular covalent crosslinking to the loss of mechanical resilience in some human keratin diseases. These insights will also provide inspiration for the synthesis of new keratin-based biomaterials.

Polymers 14(3), 614 (2022)   DOI: 10.3390/polym14030614  

 

F Sauer, A Fritsch, S Grosser, S Pawlizak, T Kießling, M Reiss-Zimmermann, M Shahryari, WC Müller, K-T Hoffmann, JA Käs, I Sack:

Whole tissue and single cell mechanics are correlated in human brain tumors Abstract:Biomechanical changes are critical for cancer progression. However, the relationship between the rheology of single cells measured ex-vivo and the living tumor is not yet understood. Here, we combined single-cell rheology of cells isolated from primary tumors with in vivo bulk tumor rheology in patients with brain tumors. Eight brain tumors (3 glioblastoma, 3 meningioma, 1 astrocytoma, 1 metastasis) were investigated in vivo by magnetic resonance elastography (MRE), and after surgery by the optical stretcher (OS). MRE was performed in a 3-Tesla clinical MRI scanner and magnitude modulus |G*|, loss angle φ, storage modulus G', and loss modulus G'' were derived. OS experiments measured cellular creep deformation in response to laser-induced step stresses. We used a Kelvin-Voigt model to deduce two parameters related to cellular stiffness (μKV) and cellular viscosity (ηKV) from OS measurements in a time regimen that overlaps with that of MRE. We found that single-cell μKV was correlated with |G*| (R = 0.962, p < 0.001) and G'' (R = 0.883, p = 0.004) but not G' of the bulk tissue. These results suggest that single-cell stiffness affects tissue viscosity in brain tumors. The observation that viscosity parameters of individual cells and bulk tissue were not correlated suggests that collective mechanical interactions (i.e. emergent effects or cellular unjamming) of many cancer cells, which depend on cellular stiffness, influence the mechanical dissipation behavior of the bulk tissue. Our results are important to understand the emergent rheology of active multiscale compound materials such as brain tumors and its role in disease progression.

Soft Matter, 17, 10744 - 10752 (2021)   DOI: 10.1039/D1SM01291F  

 

E Warmt, S Grosser, E Blauth, X Xie, R Stange, J Tomm, F Sauer, J Schnauß, M von Bergen, JA Käs:

Differences in cortical contractility between healthy epithelial and cancerous mesenchymal breast cells Abstract:Cell contractility is mainly imagined as a force dipole-like interaction based on actin stress fibers that pull on cellular adhesionsites. Here, we present a different type of contractility based on isotropic contractions within the actomyosin cortex. Measuring mechanosensitive cortical contractility of suspended cells among various cell lines allowed us to exclude effects caused by stressfibers. We found that epithelial cells display a higher cortical tension than mesenchymal cells, directly contrasting to stressfiber-mediated contractility. These two types of contractility can even be used to distinguish epithelial from mesenchymal cells.These findings from a single cell level correlate to the rearrangement effects of actomyosin cortices within cells assembled inmulticellular aggregates. Epithelial cells form a collective contractile actin cortex surrounding multicellular aggregates and further generate a high surface tension reminiscent of tissue boundaries. Hence, we suggest this intercellular structure as tobe crucial for epithelial tissue integrity. In contrast, mesenchymal cells do not form collective actomyosin cortices reducing multicellular cohesion and enabling cell escape from the aggregates.

New Journal of Physics, 23, 103020 (2021),   DOI: 10.1088/1367-2630/ac254e  

 

E Blauth, H Kubitschke, P Gottheil, S Grosser, JA Käs:

Jamming in Embryogenesis and Cancer Progression Abstract:The ability of tissues and cells to move and rearrange is central to a broad range of diverse biological processes such as tissue remodeling and rearrangement in embryogenesis, cell migration in wound healing, or cancer progression. These processes are linked to a solid-like to fluid-like transition, also known as unjamming transition, a not rigorously defined framework that describes switching between a stable, resting state and an active, moving state. Various mechanisms, that is, proliferation and motility, are critical drivers for the (un)jamming transition on the cellular scale. However, beyond the scope of these fundamental mechanisms of cells, a unifying understanding remains to be established. During embryogenesis, the proliferation rate of cells is high, and the number density is continuously increasing, which indicates number-density-driven jamming. In contrast, cells have to unjam in tissues that are already densely packed during tumor progression, pointing toward a shape-driven unjamming transition. Here, we review recent investigations of jamming transitions during embryogenesis and cancer progression and pursue the question of how they might be interlinked. We discuss the role of density and shape during the jamming transition and the different biological factors driving it.

Frontiers in Physics, 17 August (2021),   DOI: 10.3389/fphy.2021.666709  

 

J Schnauß, T Kunschmann, S Grosser, P Mollenkopf, T Zech, JS Freitag, D Prascevic, R Stange, LS Röttger, S Rönicke, DM Smith, TM Bayerl, JA Käs:

Cells in Slow Motion: Apparent Undercooling Increases Glassy Behavior at Physiological Temperatures Abstract:Solvent conditions are unexpectedly sufficient to drastically and reversibly slow down cells. In vitro on the molecular level, protein-solvent interactions drastically change in the presence of heavy water (D2O) and its stronger hydrogen bonds. Adding D2O to the cell medium of living cells increases the molecular intracellular viscosity. While cell morphology and phenotype remain unchanged, cellular dynamics transform into slow motion in a changeable manner. This is exemplified in the slowdown of cell proliferation and migration, which is caused by a reversible gelation of the cytoplasm. In analogy to the time-temperature superposition principle, where temperature is replaced by D2O, an increase in viscosity slows down the effective time. Actin networks, crucial structures in the cytoplasm, switch from a power-law-like viscoelastic to a more rubber-like elastic behavior. The resulting intracellular resistance and dissipation impair cell movement. Since cells are highly adaptive non-equilibrium systems, they usually respond irreversibly from a thermodynamic perspective. D2O induced changes, however, are fully reversible and their effects are independent of signaling as well as expression. The stronger hydrogen bonds lead to glass-like, drawn-out intramolecular dynamics, which may facilitate longer storage times of biological matter, for instance, during transport of organ transplants.

Advanced Materials 2101840 (2021),   DOI: 10.1002/adma.202101840  

 

T Händler, C Tutmarc, M Glaser, JS Freitag, DM Smith, J Schnauß:

Measuring structural parameters of crosslinked and entangled semiflexible polymer networks with single-filament tracing Abstract:Single-filament tracing has been a valuable tool to directly determine geometrical and mechanical properties of entangled polymer networks. However, systematically verifying how the stiffness of the tracer filament or its molecular interactions with the surrounding network impacts the measurement of these parameters has not been possible with the established experimental systems. Here, we use mechanically programmable DNA nanotubes embedded in crosslinked and entangled F-actin networks, as well as in synthetic DNA networks, in order to measure fundamental, structural network properties like tube width and mesh size with respect to the stiffness of the tracers. While we confirm some predictions derived from models based purely on steric interactions, our results indicate that these models should be expanded to account for additional inter-filament interactions, thus describing the behavior of real polymer networks.

Physical Review E, accepted (2021),  

 

B Mages, T Fuhs, S Aleithe, A Blietz, C Hobusch, W Härtig, S Schob, M Krueger, D Michalski:

The Cytoskeletal Elements MAP2 and NF-L Show Substantial Alterations in Different Stroke Models While Elevated Serum Levels Highlight Especially MAP2 as a Sensitive Biomarker in Stroke Patients Abstract:In the setting of ischemic stroke, the neurofilament subunit NF-L and the microtubule-associated protein MAP2 have proven to be exceptionally ischemia-sensitive elements of the neuronal cytoskeleton. Since alterations of the cytoskeleton have been linked to the transition from reversible to irreversible tissue damage, the present study investigates underlying time- and region-specific alterations of NF-L and MAP2 in different animal models of focal cerebral ischemia. Although NF-L is increasingly established as a clinical stroke biomarker, MAP2 serum measurements after stroke are still lacking. Therefore, the present study further compares serum levels of MAP2 with NF-L in stroke patients. In the applied animal models, MAP2-related immunofluorescence intensities were decreased in ischemic areas, whereas the abundance of NF-L degradation products accounted for an increase of NF-L-related immunofluorescence intensity. Accordingly, Western blot analyses of ischemic areas revealed decreased protein levels of both MAP2 and NF-L. The cytoskeletal alterations are further reflected at an ultrastructural level as indicated by a significant reduction of detectable neurofilaments in cortical axons of ischemia-affected areas. Moreover, atomic force microscopy measurements confirmed altered mechanical properties as indicated by a decreased elastic strength in ischemia-affected tissue. In addition to the results from the animal models, stroke patients exhibited significantly elevated serum levels of MAP2, which increased with infarct size, whereas serum levels of NF-L did not differ significantly. Thus, MAP2 appears to be a more sensitive stroke biomarker than NF-L, especially for early neuronal damage. This perspective is strengthened by the results from the animal models, showing MAP2-related alterations at earlier time points compared to NF-L. The profound ischemia-induced alterations further qualify both cytoskeletal elements as promising targets for neuroprotective therapies.

Molecular Neurobiology (2021),   DOI: 10.1007/s12035-021-02372-3  

 

I Nel, EW Morawetz, D Tschodu, JA Käs, B Aktas:

The Mechanical Fingerprint of Circulating Tumor Cells (CTCs) in Breast Cancer Patients Abstract:Circulating tumor cells (CTCs) are a potential predictive surrogate marker for disease monitoring. Due to the sparse knowledge about their phenotype and its changes during cancer progression and treatment response, CTC isolation remains challenging. Here we focused on the mechanical characterization of circulating non-hematopoietic cells from breast cancer patients to evaluate its utility for CTC detection. For proof of premise, we used healthy peripheral blood mononuclear cells (PBMCs), human MDA-MB 231 breast cancer cells and human HL-60 leukemia cells to create a CTC model system. For translational experiments CD45 negative cells - possible CTCs - were isolated from blood samples of patients with mamma carcinoma. Cells were mechanically characterized in the optical stretcher (OS). Active and passive cell mechanical data were related with physiological descriptors by a random forest (RF) classifier to identify cell type specific properties. Cancer cells were well distinguishable from PBMC in cell line tests. Analysis of clinical samples revealed that in PBMC the elliptic deformation was significantly increased compared to nonhematopoietic cells. Interestingly, non-hematopoietic cells showed significantly higher shape restoration. Based on Kelvin-Voigt modeling, the RF algorithm revealed that elliptic deformation and shape restoration were crucial parameters and that the OS discriminated non-hematopoietic cells from PBMC with an accuracy of 0.69, a sensitivity of 0.74, and specificity of 0.63. The CD45 negative cell population in the blood of breast cancer patients is mechanically distinguishable from healthy PBMC. Together with cell morphology, the mechanical fingerprint might be an appropriate tool for marker-free CTC detection.

Cancers, 13 (5), 1119 (2021),   DOI: 10.3390/cancers13051119  

 

S Grosser, J Lippoldt, L Oswald, M Merkel, DM Sussman, F Renner, P Gottheil, EW Morawetz, T Fuhs, X Xie, S Pawlizak, AW Fritsch, B Wolf, L-C Horn, S Briest, B Aktas, ML Manning, JA Käs:

Cell and Nucleus Shape as an Indicator of Tissue Fluidity in Carcinoma Abstract:Tissue, cell, and nucleus morphology change during tumor progression. In 2D confluent cell cultures, different tissue states, such as fluid (unjammed) and solid (jammed) are correlated with cell shapes. These results do not have to apply a priori to three dimensions. Cancer cell motility requires and corresponds to a fluidization of the tumor tissue on the bulk level. Here we investigate bulk tissue fluidity in 3D and determine how it correlates with cell and nucleus shape. In patient samples of mamma and cervix carcinoma we find areas where cells can move or are immobile. We compare 3D cell spheroids composed of cells from a cancerous and a non-cancerous cell line. Through bulk mechanical spheroid-fusion experiments and single live-cell tracking, we show that the cancerous sample is fluidized by active cells moving through the tissue. The healthy, epithelial sample with immobile cells behaves more solid-like. 3D-segmentations of the samples show that the the degree of tissue fluidity correlates with elongated cell and nucleus shapes. This links cell shapes to cell motility and bulk mechanical behavior. We find two active states of matter in solid tumors: an amorphous glass-like state with characteristics of 3D cell jamming, and a disordered fluid state. Individual cell and nucleus shape may serve as marker for metastatic potential to foster personalized cancer treatment.

Physical Review X 11, 011033 (2021),   DOI: 10.1103/PhysRevX.11.011033  

 

O Ilina, PG Gritsenko, S Syga, J Lippoldt, CAM La Porta, O Chepizhko, S Grosser, M Vullings, GJ Bakker, J Starruß, P Bult, S Zapperi, JA Käs, A Deutsch, P Friedl:

Cell-Cell Adhesion and 3D Matrix Confinement Determine Jamming Transitions in Breast Cancer Invasion Abstract:Plasticity of cancer invasion and metastasis depends on the ability of cancer cells to switch between collective and single-cell dissemination, controlled by cadherin-mediated cell-cell junctions. In clinical samples, E-cadherin-expressing and -deficient tumours both invade collectively and metastasize equally, implicating additional mechanisms controlling cell-cell cooperation and individualization. Here, using spatially defined organotypic culture, intravital microscopy of mammary tumours in mice and in silico modelling, we identify cell density regulation by three-dimensional tissue boundaries to physically control collective movement irrespective of the composition and stability of cell-cell junctions. Deregulation of adherens junctions by downregulation of E-cadherin and p120-catenin resulted in a transition from coordinated to uncoordinated collective movement along extracellular boundaries, whereas single-cell escape depended on locally free tissue space. These results indicate that cadherins and extracellular matrix confinement cooperate to determine unjamming transitions and stepwise epithelial fluidization towards, ultimately, cell individualization.

Nature Cell Biology 22, 1103 - 1115 (2020),   DOI: 10.1038/s41556-020-0552-6  

 

J Schnauß, BUS Schmidt, CB Brazel, S Dogan, W Losert, U Anderegg, JA Käs:

Influence of Hyaluronic Acid Binding on the Actin Cortex measured by Optical Forces Abstract:Melanoma cells are often surrounded by hyaluronic acid (HA) rich environments, which are considered to promote tumor progression and metastasis. Induced effects in compound materials consisting of cells embedded in an extracellular matrix (ECM) have been studied, however, alterations of the single cells have never been addressed. Here, we explicitly addressed single cell properties and measured HA-induced biomechanical changes via deformations induced solely by optical forces. With the optical stretcher setup, cells were deformed after culturing them either in the presence or absence of HA revealing the crucial interplay of HA with the CD44 receptor. To assess the role of CD44 in transducing effects of HA, we compared a CD44 expressing variant of the melanoma cell line RPM-MC to its natural CD44-negative counterpart. Our measurements revealed a significant stiffness change, which we attribute to changes of the actin cytoskeleton.

Journal of Biophotonics, e201960215 (2020),   DOI: 10.1002/jbio.20190215  

 

KJ Streitberger, L Lilaj, F Schrank, J Braun, KT Hoffmann, M Reiss-Zimmermann, JA Käs, I Sack:

How tissue fluidity influences brain tumor progression Abstract:Mechanical properties of biological tissues and, above all, their solid or fluid behavior influence the spread of malignant tumors. While it is known that solid tumors tend to have higher mechanical rigidity, allowing them to aggressively invade and spread in solid surrounding healthy tissue, it is unknown how softer tumors can grow within a more rigid environment such as the brain. Here, we use in vivo magnetic resonance elastography (MRE) to elucidate the role of anomalous fluidity for the invasive growth of soft brain tumors, showing that aggressive glioblastomas (GBMs) have higher water content while behaving like solids. Conversely, our data show that benign meningiomas (MENs), which contain less water than brain tissue, are characterized by fluid-like behavior. The fact that the 2 tumor entities do not differ in their soft properties suggests that fluidity plays an important role for a tumor's aggressiveness and infiltrative potential. Using tissue-mimicking phantoms, we show that the anomalous fluidity of neurotumors physically enables GBMs to penetrate surrounding tissue, a phenomenon similar to Saffman-Taylor viscous-fingering instabilities, which occur at moving interfaces between fluids of different viscosity. Thus, targeting tissue fluidity of malignant tumors might open horizons for the diagnosis and treatment of cancer.

PNAS January 7, 117 (1) 128 - 134 (2020),   DOI: 10.1073/pnas.1913511116  

 

P van Liedekerke, J Neitsch, T Johann, E Warmt, IG Valverde, S Grosser, S Hoehme, JA Käs, D Drasdo:

A quantitative high-resolution computational mechanics cell model for growing and regenerating tissues Abstract:Mathematical models are increasingly designed to guide experiments in biology, biotechnology, as well as to assist in medical decision making. They are in particular important to understand emergent collective cell behavior. For this purpose, the models, despite still abstractions of reality, need to be quantitative in all aspects relevant for the question of interest. This paper considers as showcase example the regeneration of liver after drug-induced depletion of hepatocytes, in which the surviving and dividing hepatocytes must squeeze in between the blood vessels of a network to refill the emerged lesions. Here, the cellsí response to mechanical stress might significantly impact the regeneration process. We present a 3D high-resolution cell-based model integrating information from measurements in order to obtain a refined and quantitative understanding of the impact of cell-biomechanical effects on the closure of drug-induced lesions in liver. Our model represents each cell individually and is constructed by a discrete, physically scalable network of viscoelastic elements, capable of mimicking realistic cell deformation and supplying information at subcellular scales. The cells have the capability to migrate, grow, and divide, and the nature and parameters of their mechanical elements can be inferred from comparisons with optical stretcher experiments. Due to triangulation of the cell surface, interactions of cells with arbitrarily shaped (triangulated) structures such as blood vessels can be captured naturally. Comparing our simulations with those of so-called center-based models, in which cells have a largely rigid shape and forces are exerted between cell centers, we find that the migration forces a cell needs to exert on its environment to close a tissue lesion, is much smaller than predicted by center-based models. To stress generality of the approach, the liver simulations were complemented by monolayer and multicellular spheroid growth simulations. In summary, our model can give quantitative insight in many tissue organization processes, permits hypothesis testing in silico, and guide experiments in situations in which cell mechanics is considered important.

Biomechanics and Modelling in Mechanobiology, 19, 189 - 220 (2020),   DOI: 10.1007/s10237-019-01204-7  

 

Y Shen, BUS Schmidt, H Kubitschke, EW Morawetz, B Wolf, JA Käs, W Losert:

Detecting heterogeneity in and between breast cancer cell lines Abstract:Cellular heterogeneity in tumor cells is a well-established phenomenon. Genetic and phenotypic cell-to-cell variability have been observed in numerous studies both within the same type of cancer cells and across different types of cancers. Another known fact for metastatic tumor cells is that they tend to be softer than their normal or non-metastatic counterparts. However, the heterogeneity of mechanical properties in tumor cells are not widely studied. Here we analyzed single-cell optical stretcher data with machine learning algorithms on three different breast tumor cell lines and show that similar heterogeneity can also be seen in mechanical properties of cells both within and between breast tumor cell lines. We identified two clusters within MDA-MB-231 cells, with cells in one cluster being softer than in the other. In addition, we show that MDA-MB-231 cells and MDA-MB-436 cells which are both epithelial breast cancer cell lines with a mesenchymal-like phenotype derived from metastatic cancers are mechanically more different from each other than from non-malignant epithelial MCF-10A cells. Since stiffness of tumor cells can be an indicator of metastatic potential, this result suggests that metastatic abilities could vary within the same monoclonal tumor cell line.

Cancer Convergence 4, Article number 1 (2020),   DOI: 10.1186/s41236-020-0010-1  

 

P M Shahryari, H Tzschätzsch, J Guo, SR Marticorena Garcia, G Böning, U Fehrenbach, L Stencel, P Asbach, B Hamm, JA Käs, J Braun, T Denecke, I Sack:

Tomoelastography Distinguishes Noninvasively between Benign and Malignant Liver Lesions Abstract:Patients with increased liver stiffness have a higher risk of developing cancer, however, the role of fluid-solid tissue interactions and their contribution to liver tumor malignancy remains elusive. Tomoelastography is a novel imaging method for mapping quantitatively the solid-fluid tissue properties of soft tissues in vivo. It provides high resolution and thus has clear clinical applications. In this work we used tomoelastography in 77 participants, with a total of 141 focal liver lesions of different etiologies, to investigate the contributions of tissue stiffness and fluidity to the malignancy of liver tumors. Shear-wave speed (c) as surrogate for tissue stiffness and phase-angle (f) of the complex shear modulus reflecting tissue fluidity were abnormally high in malignant tumors and allowed them to be distinguished from nontumorous liver tissue with high accuracy [c: AUC = 0.88 with 95% confidence interval (CI) = 0.83-0.94; f: AUC = 0.95, 95% CI = 0.92-0.98]. Benign focal nodular hyperplasia and hepatocellular adenoma could be distinguished from malignant lesions on the basis of tumor stiffness (AUC = 0.85, 95% CI = 0.72-0.98; sensitivity = 94%, 95% CI = 89-100; and specificity = 85%, 95% CI = 62-100), tumor fluidity (AUC = 0.86, 95% CI = 0.77-0.96; sensitivity = 83%, 95% CI = 72-93; and specificity = 92%, 95% CI = 77-100) and liver stiffness (AUC = 0.84, 95% CI = 0.74-0.94; sensitivity = 72%, 95% CI = 59-83; and specificity = 88%, 95% CI = 69-100), but not on the basis of liver fluidity. Together, hepatic malignancies are characterized by stiff, yet fluid tissue properties, whereas surrounding nontumorous tissue is dominated by solid properties. Tomoelastography can inform noninvasively on the malignancy of suspicious liver lesions by differentiating between benign and malignant lesions with high sensitivity based on stiffness and with high specificity based on fluidity.

Cancer Research vol. 79 iss. 22 (2019),   DOI: 10.1158/0008-5472.CAN-19-2150  

 

H Kubitschke, B Wolf, E Morawetz, L-C Horn, B Aktas, U Behn, M Höckel, JA Käs:

Roadmap to Local Tumour Growth: Insights from Cervical Cancer Abstract:Wide tumour excision is currently the standard approach to surgical treatment of solid cancers including carcinomas of the lower genital tract. This strategy is based on the premise that tumours exhibit isotropic growth potential. We reviewed and analysed local tumour spreading patterns in 518 patients with cancer of the uterine cervix who underwent surgical tumour resection. Based on data obtained from pathological examination of the surgical specimen, we applied computational modelling techniques to simulate local tumour spread in order to identify parameters influencing preferred infiltration patterns and used area-proportional Euler diagrams to detect and confirm ordered patterns of tumour spread. Some anatomical structures, e.g. tissues of the urinary bladder, were significantly more likely to be infiltrated than other structures, e.g. the ureter and the rectum. Computational models assuming isotropic growth could not explain these infiltration patterns. Introducing ontogenetic distance of a tissue relative to the uterine cervix as a parameter led to accurate predictions of the clinically observed infiltration likelihoods. The clinical data indicates that successive infiltration likelihoods of ontogenetically distant tissues are nearly perfect subsets of ontogenetically closer tissues. The prevailing assumption of isotropic tumour extension has significant shortcomings in the case of cervical cancer. Rather, cervical cancer spread seems to follow ontogenetically defined trajectories.

Scientific Reports 9, Article number 12768 (2019),   DOI: 10.1038/s41598-019-49182-1  

 

C Ficorella, R Martinez Vàzques, P Heine, E Lepera, J Cao, E Warmt, R Osellame, JA Käs:

Normal epithelial and triple-negative breast cancer cells show the same invasion potential in rigid spatial confinement Abstract:The extra-cellular microenvironment has a fundamental role in tumor growth and progression, strongly affecting the migration strategies adopted by single cancer cells during metastatic invasion. In this study, we use a novel microfluidic device to investigate the ability of mesenchymal and epithelial breast tumor cells to fluidize and migrate through narrowing microstructures upon chemoattractant stimulation. We compare the migration behavior of two mesenchymal breast cancer cell lines and one epithelial cell line, and find that the epithelial cells are able to migrate through the narrowest microconstrictions as the more invasive mesenchymal cells. In addition, we demonstrate that migration of epithelial cells through a highly compressive environment can occur in absence of a chemoattractive stimulus, thus evidencing that they are just as prone to react to mechanical cues as invasive cells.

New Journal of Physics, 21 (2019),   DOI: 10.1088/1367-2630/ab3572  

 

T Golde, M Glaser, C Tutmarc, I Elbalasy, C Huster, G Busteros, DM Smith, H Herrmann, JA Käs, J Schnauß:

The role of stickiness in the rheology of semiflexible polymers Abstract:Semiflexible polymers form central structures in biological material. Modelling approaches usually neglect influences of polymer-specific molecular features aiming to describe semiflexible polymers universally. Here, we investigate the influence of molecular details on networks assembled from filamentous actin, intermediate filaments, and synthetic DNA nanotubes. In contrast to prevalent theoretical assumptions, we find that bulk properties are affected by various inter-filament interactions. We present evidence that these interactions can be merged into a single parameter in the frame of the glassy wormlike chain model. The interpretation of this parameter as a polymer specific stickiness is consistent with observations from macro-rheological measurements and reptation behaviour. Our findings demonstrate that stickiness should generally not be ignored in semiflexible polymer models.

Soft Matter, 15, 4865-4872 (2019),   DOI: 10.1039/C9SM00433E 

 

F Sauer, L Oswald, A Ariza de Schellenberger, H Tzschätzsch, F Schrank, T Fischer, J Braun, CT Mierke, R Valiullin, I Sack, JA Käs:

Collagen networks determine viscoelastic properties of connective tissues yet do not hinder diffusion of the aqueous solvent Abstract:Collagen accounts for the major extracellular matrix (ECM) component in many tissues and provides mechanical support for cells. Magnetic Resonance (MR) Imaging, MR based diffusion measurements and MR Elastography (MRE) are considered sensitive to the microstructure of tissues including collagen networks of the ECM. However, little is known whether water diffusion interacts with viscoelastic properties of tissues. This study combines highfield MR based diffusion measurements, novel compact tabletop MRE and confocal microscopy in collagen networks of different cross-linking states (untreated collagen gels versus additional treatment with glutaraldehyde). The consistency of bulk rheology and MRE within a wide dynamic range is demonstrated in heparin gels, a viscoelastic standard for MRE. Additional crosslinking of collagen led to an 8-fold increased storage modulus, a 4-fold increased loss modulus and a significantly decreased power law exponent, describing multi-relaxational behavior, corresponding to a pronounced transition from viscous-soft to elastic-rigid properties. Collagen network changes were not detectable by MR based diffusion measurements and microscopy which are sensitive to the micrometer scale. The MRE-measured shear modulus is sensitive to collagen fiber interactions which take place on the intrafiber level such as fiber stiffness. The insensitivity of MR based diffusion measurements to collagen hydrogels of different cross-linking states alludes that congeneric collagen structures in connective tissues do not hinder extracellular diffusive water transport. Furthermore, the glutaraldehyde induced rigorous changes in viscoelastic properties indicate that intrafibrillar dissipation is the dominant mode of viscous dissipation in collagen-dominated connective tissue.

Soft Matter, 15, 3055-3064 (2019),   DOI: 10.1039/C8SM02264J 

 

CT Mierke, F Sauer, S Grosser, S Puder, T Fischer, JA Käs:

The two faces of enhanced stroma: Stroma acts as a tumor promoter and a steric obstacle Abstract: In addition to genetic, morphological and biochemical alterations in cells, a key feature of the malignant progression of cancer is the stroma, including cancer cell motility as well as the emergence of metastases. Our current knowledge with regard to the biophysically driven experimental approaches of cancer progression indicates that mechanical aberrations are major contributors to the malignant progression of cancer. In particular, the mechanical probing of the stroma is of great interest. However, the impact of the tumor stroma on cellular motility, and hence the metastatic cascade leading to the malignant progression of cancer, is controversial as there are two different and opposing effects within the stroma. On the one hand, the stroma can promote and enhance the proliferation, survival and migration of cancer cells through mechanotransduction processes evoked by fiber alignment as a result of increased stroma rigidity. This enables all types of cancer to overcome restrictive biological capabilities. On the other hand, as a result of its structural constraints, the stroma acts as a steric obstacle for cancer cell motility in dense three-dimensional extracellular matrices, when the pore size is smaller than the cell´s nucleus. The mechanical properties of the stroma, such as the tissue matrix stiffness and the entire architectural network of the stroma, are the major players in providing the optimal environment for cancer cell migration. Thus, biophysical methods determining the mechanical properties of the stroma, such as magnetic resonance elastography, are critical for the diagnosis and prediction of early cancer stages. Fibrogenesis and cancer are tightly connected, as there is an elevated risk of cancer on cystic fibrosis or, subsequently, cirrhosis. This also applies to the subsequent metastatic process.

NMR Biomed., vol. 10, e3831 (2018),  DOI: 10.1002/nbm.3831

 

T Golde, C Huster, M Glaser, T Händler, H Herrmann, JA Käs, J Schnauß:

Glassy dynamics in composite biopolymer networks Abstract: The cytoskeleton is a highly interconnected meshwork of strongly coupled subsystems providing mechanical stability as well as dynamic functions to cells. To elucidate the underlying biophysical principles, it is central to investigate not only one distinct functional subsystem but rather their interplay as composite biopolymeric structures. Two of the key cytoskeletal elements are actin and vimentin filaments. Here, we show that composite networks reconstituted from actin and vimentin can be described by a superposition of two non-interacting scaffolds. Arising effects are demonstrated in a scale-spanning frame connecting single filament dynamics to macro-rheological network properties. The acquired results of the linear and non-linear bulk mechanics can be captured within an inelastic glassy wormlike chain model. In contrast to previous studies, we find no emergent effects in these composite networks. Thus, our study paves the way to predict the mechanics of the cytoskeleton based on the properties of its single structural components.

Soft Matter vol. 14, 7970-7978 (2018),  DOI: 10.1039/C8SM01061G

 

F Meinhövel, R Stange, J Schnauß, M Sauer, JA Käs, TW Remmerbach:

Changing cell mechanics - a precondition for malignant transformation of oral squamous carcinoma cells Abstract: Oral squamous cell carcinomas (OSCC) are the sixth most common cancer and the diagnosis is often belated for a curative treatment. The reliable and early differentiation between healthy and diseased cells is the main aim of this study in order to improve the quality of the treatment and to understand tumour pathogenesis. Here, the optical stretcher is used to analyse mechanical properties of cells and their potential to serve as a marker for malignancy. Stretching experiments revealed for the first time that cells of primary OSCCs were deformed by 2.9% rendering them softer than cells of healthy mucosa which were deformed only by 1.9%. Furthermore, the relaxation behaviour of the cells revealed that these malignant cells exhibit a faster contraction than their benign counterparts. This suggests that deformability as well as relaxation behaviour can be used as distinct parameters to evaluate emerging differences between these benign and malignant cells. Since many studies in cancer research are performed with cancer cell lines rather than primary cells, we have compared the deformability and relaxation of both types, showing that long time culturing leads to softening of cells. The higher degree of deformability and relaxation behaviour can enable cancer cells to traverse tissue emphasizing that changes in cell architecture may be a potential precondition for malignant transformation. Respecting the fact that even short culture times have an essential effect on the significance of the results, the use of primary cells for further research is recommended. The distinction between malignant and benign cells would enable an early confirmation of cancer diagnoses by testing cell samples of suspect oral lesions.

Convergent Science Physical Oncology, vol. 4, 034001(2018),  DOI: 10.1088/2057-1739/aac72d

 

J Lorenz, J Schnauß, M Glaser, M Sajfutdinow, C Schuldt, JA Käs, DM Smith:

Synthetic Transient Crosslinks Program the Mechanics of Soft, Biopolymer-Based Materials Abstract: Actin networks are adaptive materials enabling dynamic and static functions of living cells. A central element for tuning their underlying structural and mechanical properties is the ability to reversibly connect, i.e., transiently crosslink, filaments within the networks. Natural crosslinkers, however, vary across many parameters. Therefore, systematically studying the impact of their fundamental properties like size and binding strength is unfeasible since their structural parameters cannot be independently tuned. Herein, this problem is circumvented by employing a modular strategy to construct purely synthetic actin crosslinkers from DNA and peptides. These crosslinkers mimic both intuitive and noncanonical mechanical properties of their natural counterparts. By isolating binding affinity as the primary control parameter, effects on structural and dynamic behaviors of actin networks are characterized. A concentration-dependent triphasic behavior arises from both strong and weak crosslinkers due to emergent structural polymorphism. Beyond a certain threshold, strong binding leads to a nonmonotonic elastic pulse, which is a consequence of self-destruction of the mechanical structure of the underlying network. The modular design also facilitates an orthogonal regulatory mechanism based on enzymatic cleaving. This approach can be used to guide the rational design of further biomimetic components for programmable modulation of the properties of biomaterials and cells.

Advanced Materials 1706092 (2018),  DOI: 10.1002/adma.201706092

 

L Oswald, S Grosser, DM Smith, JA Käs:

Jamming transitions in cancer Abstract: The traditional picture of tissues, where they are treated as liquids defined by properties such as surface tension or viscosity has been redefined during the last few decades by the more fundamental question: Under which conditions do tissues display liquid-like or solid-like behaviour? As a result, basic concepts arising from the treatment of tissues as solid matter, such as cellular jamming and glassy tissues, have shifted into the current focus of biophysical research. Here, we review recent works examining the phase states of tissue with an emphasis on jamming transitions in cancer. When metastasis occurs, cells gain the ability to leave the primary tumour and infiltrate other parts of the body. Recent studies have shown that a linkage between an unjamming transition and tumour progression indeed exists, which could be of importance when designing surgery and treatment approaches for cancer patients.

Journal of Physics D: Applied Physics, Volume 50, 483001 (2017),  DOI: 10.1088/1361-6463/aa8e83

 

D Strehle, P Mollenkopf, M Glaser, T Golde, C Schuldt, JA Käs, J Schnauß:

Single Actin Bundle Rheology Abstract:Bundled actin structures play an essential role in the mechanical response of the actin cytoskeleton in eukaryotic cells. Although responsible for crucial cellular processes, they are rarely investigated in comparison to single filaments and isotropic networks. Presenting a highly anisotropic structure, the determination of the mechanical properties of individual bundles was previously achieved through passive approaches observing bending deformations induced by thermal fluctuations. We present a new method to determine the bending stiffness of individual bundles, by measuring the decay of an actively induced oscillation. This approach allows us to systematically test anisotropic, bundled structures. Our experiments revealed that thin, depletion force-induced bundles behave as semiflexible polymers and obey the theoretical predictions determined by the wormlike chain model. Thickening an individual bundle by merging it with other bundles enabled us to study effects that are solely based on the number of involved filaments. These thicker bundles showed a frequency-dependent bending stiffness, a behavior that is inconsistent with the predictions of the wormlike chain model. We attribute this effect to internal processes and give a possible explanation with regard to the wormlike bundle theory.

Molecules 22(10) 1804 (2017),   DOI: 10.3390/molecules22101804