Genetic transformations, biochemical communications and physical interactions are interconnected processes involved from the initial steps of tumor formation until the latter phases of cancer metastasis, a major cause of cancer related deaths. Metastatic cascade begins when the primary tumor cells gain aggressive and migratory phenotypes resulting in leaving the primary tumor (Figure 1A), invading the local tissue (Figure 1B) (Craene and Berx, 2013) and transmigrating through the endothelial barrier into the blood or lymphatic microvasculature (a process known as intravasation, Figure 1C) (Van Zijl et al., 2011). The latter steps of disease involve survival of cancer cells in blood circulation (Figure 1D) (Aceto et al., 2015; Reymond et al., 2013) and exit from the vessels at distal tissues (a process known as extravasation) to ultimately invade and colonize in the secondary sites (Figure 1E) (Nguyen et al., 2009). In addition to a number of unique genetic and biochemical factors associated with metastasis (Suresh, 2007; Kumar and Weaver, 2009; Wirtz et al., 2011), irregular mechanical alterations such as structural, morphological and stiffness changes, in both cells and the extracellular environment, play a significant role during all stages of cancer initiation and dissemination. For example, the primary tumor is characterized by biochemically and mechanically altered environment that results from oncogenic mutations and epigenetic changes disrupting key physiological cellular processes such as cell cycle (Spill et al., 2016). Taking advantage of the abnormal mechanical properties of most tumors, palpation has been a conventional diagnostic method to assess the stiffness of tumor within the surrounding soft tissue. The disease stage is linked with tumor stiffness as monitored by in vivo MRI elastography or through ex vivo atomic force microscopy (Lopez et al., 2011). On the other hand, during tumor growth, the mechanical alterations in tumor environments trigger complex bio-mechanical signalling pathways, which may ultimately enhance the ability of tumor cells to acquire a malignant phenotype. The malignant evolution of cancer cells de-regulates cell-cell and cell-extracellular matrix (ECM) adhesions and cytoskeletal remodeling leading to abnormal tumor cell morphology, enhancement of metabolism (Torrino et al., 2021), and migratory behavior which facilitate invasion at the primary site and ultimately leads to intravasation. Following intravasation, circulating tumor cells (CTCs) must resist the mechanical forces in the bloodstream to survive and reach the secondary organ. Once at the secondary site, tumor cells must exert forces and undergo morphological changes to escape from the vasculature and invade the ECM of the distal organ.

These mechanical interactions during the metastatic journey of tumor cells highlight the importance of biomechanics in cancer dissemination (Kumar and Weaver, 2009; Moeendarbary and Harris, 2014; Malandrino et al., 2018a). Here we describe the crucial interplay between mechanics and biology during cancer initiation and progression together with recent conceptual and technological advances in mechanobiology which have led to a remarkable progress in leveraging cancer biomechanics to develop novel therapeutic strategies.

Even in a seemingly static tissue, the extracellular microenvironment exerts forces to the cells. Such forces are originated from adjacent cells, the interstitial fluid, or interaction with the ECM, and are sensed via cellular mechanosensitive receptors that regulate major cellular functions such as the cell cycle, morphogenesis, and migration (Wang et al., 1993). Indeed, the bio-mechanical interactions between intracellular biological machinery and the surrounding microenvironment, through mechanosensitive receptors, create a normal physiological condition. However, in the setting of cancer, genetic and epigenetic (Burdziak et al., 2023), including biochemical and physical, perturbations in intracellular or extracellular environments, disrupt the cellular homeostasis leading to dysplasia and in most cases to the formation of a solid tumor. During tumor growth, the aforementioned normal forces are disrupted as a result of mechanical stresses originated from aberrant homeostasis, excessive growth and tissue dysplasia (Jain et al., 2014). Tumor growth is known to generate compressional forces that perturb the interstitial space, the ECM and the flow in the vasculature. In turn, perturbations in the interstitial space may cause accumulation of growth factors and cytokines that facilitate tumor growth; whereas disruption in the mechanical properties of ECM and flow may alter cellular behaviour (Malandrino et al., 2018b).

It is now evident that the mechanical aspects such as ECM stiffness are critical in regulating a wide range of cellular behaviour. For example, in the context of stem cell differentiation, human mesenchymal stem cells preferentially differentiate into neurons or osteocytes when cultured on substrates with stiffnesses matching brain or bone tissues respectively (Saha et al., 2008). In the setting of cancer transformation, when epithelial cells are cultured on a compliant substrate, normal cells show a decrease in the rate of DNA synthesis and an increase in the rate of apoptosis while transformed cancer cells maintain their growth and apoptotic characteristics (Wang et al., 2000). Furthermore, transformed cells exert higher traction forces compared to non-transformed cells. Consequently, the increase in ECM stiffness and the extent of compression can lead to activation and increased expression of Rho GTPase and downstream effectors as well as high levels of extracellular signal-regulated kinases (ERK) activity that facilitate the process of epithelial-mesenchymal transformation (EMT) (Klein et al., 2009).

Biochemical and biophysical characteristics of the ECM influences cell migration (Barenholz-Cohen et al., 2020) through variations in growth factors or chemokines (chemotaxis), stiffness (durotaxis), ligand density (haptotaxis), and topographical organization (contact guidance) to direct cells to target destinations (Wang et al., 2019). Recent advances in intravital imaging have revealed that cells can adopt a diverse set of migration strategies involving migration as single cells or collective strands, transitions between mesenchymal, epithelial, and amoeboid migration modes, deformation of the cell body and nucleus to squeeze through matrix pores, and remodeling of matrix structure to bypass the physical barriers presented by the ECM (Wang et al., 2019) (Figure 1B). Furthermore, heterogeneity of stiffness in tumor microenvironment, triggered by matrix remodeling can mechanically guide the tumor cells directional migration (Zhang et al., 2020).

EMT is a critical process in metastasis and involves loss of epithelial characteristics (Bocci et al., 2019), resulting from downregulation of cell-cell adhesion strength (for example, through loss of E-cadherin and cytokeratin) and acquisition of a mesenchymal phenotype via activation of migratory processes (for example, through upregulation of vimentin and N-cadherin). Taken together, the EMT process disrupts cellular force balances and polarity leading to morphological changes and detachment of tumor cells from the tumor epithelium (Thiery and Sleeman, 2006; Kalluri and Weinberg, 2009; Chaffer and Weinberg, 2011). By developing a high-throughput screening assay to track displacements generated by 3D cultured multicellular clusters, Leggett et al. (2020) showed a successive reduction in protrusive and circumferential tractions during EMT. Subsequently, the modulation of cellular shape and forces in combination with mechanisms favouring migration including proteolytic (matrix metalloproteinase), adhesive, protrusive (invadopodia) and contractile processes, promote invasion of cancer cells (Wolf and Friedl, 2009). Therefore, to facilitate their three-dimensional motility, cancer cells navigate through the ECM via invadopodial protrusions, balance cell-ECM adhesion, and apply contractile forces to squeeze through ECM pores and ultimately digest and remodel the ECM via force application and matrix metalloproteinase secretion.

The generation of a new tumor-specific vasculature that aids tumor growth, is concomitant with tumor development and transformation to malignancy and facilitates the escape of tumor cells into the circulation. In addition to biochemical signals (Chen et al., 2021), physical factors such as mechanical, hydrodynamical, and collective processes (Rieger and Welter, 2015) influence the generation and architecture of tumor-specific vasculature. Indeed, the growth of an avascular tumor is limited to a critical size (<1 mm) because of the inability of diffusion mechanisms to supply oxygen and soluble factors into the tumor core. This phenomenon results in the development of a necrotic/hypoxic region at the tumor core, which is surrounded by a highly proliferative outer rim. Therefore, robust vascularization mechanisms are recruited to boost tumor growth in order to enhance delivery of different factors, such as oxygen. Vascularization of the tumor and surrounding areas is initiated and maintained through the recruitment and activation of endothelial cells, mainly triggered by hypoxic pathways (Madsen et al., 2015), and signaling with other supporting cells in the tumor microenvironment. These include, immune cells, mesenchymal stromal cells and fibroblasts (Figure 1A) (Weis and Cheresh, 2011). The newly developed vessels perturb the normal architecture of blood and lymphatic networks and induce an aberrant interaction between the fluid and solid phases within the tumor leading to high levels of the interstitial fluid pressure and the lack of gas and nutrients (Figure 1A) (Koumoutsakos et al., 2013). Due to such a chemically and mechanically disordered tumor environment, direct drug delivery to solid tumors is often inefficient (Minchinton and Tannock, 2006). Following tumor vascularization, the combination of protracted tumor cell proliferation, continuous genetic transformations, angiogenesis and activation of bio-mechanical signaling pathways promote malignancy, invasion, and metastasis (Polyak and Weinberg, 2009; Wirtz et al., 2011; Mierke, 2019; Yang et al., 2020; Dou et al., 2022).

Recent technological advances provide a better insight on the biomechanical phenomena during cancer cell dissemination by probing morphological changes, mechanical properties and force interactions between cells and the extracellular environment. While advanced light microscopy techniques such as optical super-resolution imaging offer unprecedented information about the nano-scale molecular organisation of the cell (Colin-York et al., 2019a) and its link to cellular morphology and function, they are mostly limited to isolated two-dimensional (2D) cultures in which a monolayer of cells are grown on flat plastic or glass substrates. To access high resolution tumor pathophysiology in vivo, intravital imaging has revealed some fascinating morphological changes and cell migration processes associated with invasion and intravasation (Jain et al., 2002). Three dimensional (3D) interactions among cells and the extracellular environment are unique at all stages of metastasis and cannot be recapitulated in conventional 2D cultures. On the other hand, under in vivo conditions, it is extremely difficult to fully follow the temporal evolution of 3D interactions and run parametric studies to dissect the role of different factors. Therefore, advanced 3D engineered models such as microfluidic based approaches, have been used in recapitulating key biomechanical features that are specific to each step of metastasis (Chen et al., 2017; Agrawal et al., 2022; Straehla et al., 2022; Watson et al., 2022) and to quantify tumor cell secretions at the single cell resolution over a long period of culture (Hassanzadeh-Barforoushi et al., 2020). Together, they have allowed the investigation of a variety of cellular events with high spatiotemporal resolution and under tunable environments (Malandrino et al., 2018a).

The measurement of mechanical properties is critically important in cancer biomechanics. Mechanical changes and alternations in the composition, architecture and stiffness of tumor microenvironment regulate tumor growth, transformation to malignancy, and invasion (Pickup et al., 2014) being therefore critical aspects in cancer progression (Cox and Erler, 2011; Lu et al., 2012; Popova and Jücker, 2022). For instance, tumor-associated collagen exhibits specific features due to variation in fiber orientation and collagen deposition. Notably, three distinct tumor-associated collagen signatures (TACS) have been identified in relation to human breast cancers: the accumulation of collagen fibers around small tumors (TACS-1), the straighten fibers in the vicinity of non-invasive tumors (TACS-2) and the perpendicular alignment of collagen fibers at the tumor periphery (TACS-3) (Warli et al., 2023). Such irregularities have been visualized through microscopy techniques, such as second harmonic generation, which highlights the structural transformations in collagen fibers (Figure 2A). Furthermore, aberrant mechanical properties of the tumor environment can be measured as a diagnostic tool (Plodinec et al., 2012). The mechanical properties of the tumor per se, as well as the micro-mechanical features of tumor environment have been characterized through a number of in vivo, ex vivo, and in vitro based assays at micro-nano- scale resolutions. As an alternative to conventional palpation methods, in vivo elastography measurements, such as ultrasonography (Wells and Liang, 2011), optical coherence tomography (Kennedy et al., 2015), and magnetic resonance imaging (Venkatesh et al., 2008), revealed significant stiffening of tumor tissues particularly in malignant tumors (Ramião et al., 2016; Ishihara and Haga, 2022). For example, pancreatic cancer tissue exhibits greater stiffness of ∼6 kPa compared to the 1–3 kPa range observed in normal pancreatic tissue (Itoh et al., 2016). Lung solid tumors register stiffness levels of ∼20–30 kPa, whereas normal lung parenchyma stiffness typically ranges from 0.5 to 5 kPa (Miyazawa et al., 2018). For mammary tissues, cancerous tissue displays a significantly higher stiffness of around 4 kPa, in stark contrast to the ∼0.2 kPa stiffness found in normal mammary tissue (Paszek et al., 2005). Elastography-based ultrasound techniques offer a non-invasive and real-time approach to assessing tissue stiffness, enhancing the diagnostic accuracy, and providing valuable information for personalized cancer treatment (Figure 2B). Interestingly, Golatta et al. showed that adding combined shear wave elastography and strain elastography to routine B-mode breast ultrasound could help reduce the number of unnecessary biopsies in breast diagnostics by ∼35% (Golatta et al., 2022). However, increased stiffness signature, that has been a well-known characteristic of solid tumors, alone has a limited prognostic power. The prognostic potential enhances when examining other rheological properties of the tumor microenvironment through the application of in vivo multifrequency magnetic resonance elastography (MRE). Employing this technique, Sauer et al. (2023) outlined a roadmap for prognosis of a tumor’s aggressiveness and metastatic potential based on stiffness, fluidity, spatial heterogeneity, and texture of the tumor (Figure 2C). They showed that cancer progression is accompanied by tissue fluidization, where portions of the tissue can change position across different length scales.”

Alterations in cellular and extracellular composition/structures contribute to the increased tumor stiffness. For example, excessive proliferation of cancer cells and activation of stromal cells such as cancer associated fibroblasts (CAFs) (Calvo et al., 2013), induce ECM remodeling (Bertero et al., 2019); whereas tumor growth induces solid stress within the tumor itself (Nia et al., 2017; Nia et al., 2020). The presence of growth-induced solid stresses in tumors had been under suspicion for some time, as these stresses were largely estimated using mathematical models (Stylianopoulos et al., 2012). However, in the past decade various experimental techniques have emerged to directly measure such stresses. For instance, Campàs et al. (2014) introduced incompressible oil microdroplets into 3D cell aggregates and live embryonic tissue to assess the local anisotropic forces. Dolega et al. (2017) and Lee et al. (2019) employed compressible polyacrylamide microdroplets to quantify both radial and circumferential components of solid stress within spheroids. Additionally, L et al. (2017) showed that CAFs promote tumor invasion and metastasis through exerting mechanical forces on cancer cells which are mediated via adhesion proteins involving N-cadherin at the CAF membrane and E-cadherin at the cancer cell membrane. On the other hand, tumor vascularization leads to aberrant interactions between the blood flow that infiltrates and surrounds the tumor and the increased interstitial fluid pressure (Jain et al., 2014). To dissect the contribution of the different components of tumor microenvironment, high-resolution mechanical measurement techniques, such as AFM (atomic force microscopy), have been widely applied. AFM nanomechanical indentation tests on tumor slices showed a soft mechanical signature within the tumor core, where cancer cells are abundant, while the adjacent peripheral regions are stiffened mostly due to collagen alignment (Figure 2D) (Laklai et al., 2016; Stylianou and Stylianopoulos, 2016).

At the single cell level, several experimental methods such as magnetic twisting cytometry, magnetic and optical tweezers (Figure 2E) and AFM have been utilized to perturb small regions of the cell and characterize mechanical properties, such as stiffness, of isolated cancer cells. Furthermore, Kim et al. (2018) developed a multi-parametric single-cell-analysis method in which different cell lines were transported through a microfluidics channel to measure their mechanical properties. By defining a whole-cell deformability index, they showed that malignant and non-malignant cell lines have different mechanical signatures. Traction Force Microscopy has been widely utilized to observe force interaction of cells and measure cell-generated forces (Colin-York et al., 2019b; Javanmardi et al., 2021; Li et al., 2021). This force probing technique is based on imaging the cell-induced displacement of fiducial markers, embedded/targeted within extracellular environment, and use the computational procedure to back-calculate the cellular forces that generated the displacements. To streamline and simplify the computation procedures involved in TFM, Barrasa-Fano et al. (2021) developed TFMLAB, a MATLAB software package for 4D TFM. Interestingly, while highly metastatic cancer cells exhibited a softer phenotype (∼0.5 kPa) compared to non-metastatic cells (∼2 kPa), they generate stronger forces (Kraning-Rush et al., 2012; Kristal-Muscal et al., 2013) (∼300 nN for MDA-MB231 vs ∼150 nN for MCF10A) (Cross et al., 2007; Gal and Weihs, 2012)”, allowing them to squeeze through 3D ECM and metastasize more readily (Coughlin et al., 2013). Also, in the cluster level, contractile forces generated by tumor spheroid have been measured and normalized in a scale-independent manner (Mark et al., 2020).

The mechanical behavior of ECM, cancer cells, and tissues are often assumed to be elastic solids for simplicity and their time-dependent mechanical responses are frequently overlooked. However, it has been demonstrated that ECM exhibits a more complex mechanical behavior, including viscoelasticity, mechanical plasticity, and nonlinear elasticity (Chaudhuri et al., 2020) and indeed, the viscoelasticity of ECM plays a fundamental role in the progression of cancer (Mierke, 2021). Additionally, cellular behavior is not affected by only the mechanics of solid compartment of the ECM; viscosity of the extracellular fluid has also been shown to facilitate tumor cell migration and dissemination on 2D surfaces and in 3D spheroids (Bera et al., 2022). In addition to ECM, cancer cells themselves typically show a lower levels of viscosity and membrane tension compared to healthy cells (Ren et al., 2023). Moreover, cytoplasm of the living cells also has been shown to behave as a poroelastic material (Moeendarbary et al., 2013) with an enhanced diffusion coefficient for cancer cells (Ren et al., 2023). At the tissue level, higher levels of viscos (loss) modulus in cancerous tissues (Deptuła et al., 2020) has improved the diagnostic accuracy and capabilities in ultrasound (Nabavizadeh et al., 2019) and MRE techniques (Reiter et al., 2022).

It is well known that tumor cells can vary widely according to the oncogenic background. Often, further heterogeneity occurs within a metastatic tumor when different patients are considered. Lab-on-a-chip systems are particularly well poised to address such questions related to patient -specificity with key advantages such as the requirement of low sample and reagent inputs, which are often scarce when derived directly from a patient (Figure 2F). Further, the ability to multiplex and perform high-throughput experiments is particularly amenable in lab-on-a-chip assays, enabling clinical level drug screening to be done on a person-to-person basis. For instance, microfluidic assays that recapitulate the surrounding mechanical environment of tumor cells (matrix stiffness, composition, pore size) can be used to understand the migratory phenotype of individual tumor cells as a typical indicator of potential malignancy, and how different pharmacological perturbations modulate this behavior (Pathak and Kumar, 2012; Polacheck et al., 2013; Polacheck et al., 2014) (Figure 2G). More complex in vitro models that mirror not only the surrounding ECM but also other physical cues (such as vasculature and stromal cells and fluid flows (Achilli et al., 2012; Kim et al., 2015; Whelan et al., 2023) can be employed in a multiplexed and high throughput manner to understand how these variables influence tumor cell migration in a patient-specific manner. This is particularly important as the recent FDA modernization act aims to integrate complex in vitro models of different diseases into the drug development process. For instance, Prolyl hydroxylases (PHDs) inhibitors have been shown to improve drug distribution in mice tumors and increase the effectiveness of chemotherapy. To demonstrate such effects in human cell models, Hu et al. (2022) employed a perfusable vascularized spheroid-on-a-chip model to simulate tumor microenvironment in vivo. They showed that dimethylallyl glycine improves the efficacy of the anticancer drugs paclitaxel and cisplatin in human esophageal carcinoma (Eca-109) spheroids (Figure 2F). Additionally, this could allow further understanding of the effects of external physical features such as matrix composition, stiffness, pore size, tumor cell adhesion molecules, and MMPs, on tumor migratory abilities and potentially tumor type characterization.

In addition to personalized drug screening under relevant biomechanical condition, it may also be possible phenotyping tumor cells and assess for instance their degree of malignancy through mechanical measurements on patient derived single tumour cells. Stiffness values of tumor cells or even aggregates of cells like tumor spheroids can now be measured using various contact-based (e.g., AFM) and non-contact-based methods (e.g., Brillouin imaging (Scarcelli et al., 2015; Roberts et al., 2021)); correlations can be made between these stiffness values and the invasive properties of these cells, through parallel in vitro screening experiments, and even using clinical tumor phenotyping data. This could allow extrapolating the metastatic potential from a small number of patient-derived samples and would prove to be a useful predictor when used in conjunction with traditional diagnostic methods.

Lastly, a deep understanding of the physical and structural properties of the surrounding tumor matrix might be useful for optimizing drug delivery to tumor sites. As an example, second harmonic imaging is now widely used to quantify the structural composition of collagens in the tumor microenvironment (Condeelis and Weissleder, 2010), and allow the characterization of pore size and fiber thickness, which may have a large influence on the effectiveness of drug transport. Obtaining this data for patient-specific tumors and subsequent computational modelling of drug transport based on these parameters may aid in the construction of a more optimized drug delivery system in the clinic.

Accumulated evidence has demonstrated the fundamental role of mechanobiology in cancer research as the physical properties of both cells and their microenvironment play a crucial role in the development and progression of cancer. In this short review we summarized the role of cellular biomechanical properties, e.g., stiffness, adhesion and motility, morphology, deformability, and contractility, as well as microenvironment properties such as ECM stiffness, tissue architecture, blood vessel permeability, and interstitial flow, in different steps of the tumor dissemination. Indeed, mechanobiological insight could benefit researchers and clinicians in various ways; For instance, mechanical quantification at the tissue level has been used as a diagnosis or prognosis tool for various types of cancer. Advancements in the in vivo non-invasive techniques such as MRE and ultrasound elastography as well as ex vivo techniques such as AFM have improved our diagnosis abilities in the past decade. Furthermore, at the cellular level, mechanobiology can contribute significantly to our comprehension of molecular and genetic alterations in cancer. The interplay between mechanical forces and cellular behaviors can influence how genes are expressed, proteins are synthesized, and signaling pathways are activated. Such insights are of crucial importance in the development of new therapeutic strategies, as many ongoing clinical trials target components of the extracellular matrix (Lampi and Reinhart-King, 2018; Huang et al., 2021) and proteins associated with mechanosignaling pathways (Jiang et al., 2022). Nonetheless, owing to the diversity in oncogenic backgrounds and the inherent variability within metastatic tumors, distinct therapeutic strategies are required for individual patients; In this regard, microfluidics technology holds substantial promise in advancing personalized medicine through its ability to create controlled environment for studying individualized patient samples and responses. Nevertheless, there are intriguing avenues for further research to enhance the translatability of mechanobiological knowledge derived from these platforms into clinical practice. This includes integration of the cancer mechanobiology insight with the data from genomics, transcriptomics, and proteomics approaches; this, in combination with single cell data analysis (obtained from limited number of tumor cells harvested from patient’s blood or tissue) could augment our understanding of mechanotransducive pathways involved in cancer. Moreover, by introducing immune cells along with other supporting cell types into the 3D tumor-mimicking microenvironment established within microfluidic chips, we can investigate effects of mechanical signals on immune cell behavior, as well as their implications for immunotherapy responses and the emergence of resistance.