Since physiological and pathological events change the mechanical properties of cells, tools that rapidly quantify such changes at the single-cell level can advance the utility of cell mechanics as a label-free biomarker. We demonstrate the capability to probe the population-level elastic modulus and fluidity of MDA-MB-231 cells at a throughput of up to 50 cell/second within a portable microchip. Our sensing scheme adapts a code multiplexing scheme to implement a distributed network of sensors throughout the microchip, thereby compressing all sensing events into a single electrical output. To validate our approach, we prepared cell samples whose stiffnesses were manipulated with chemical agents. We confirmed the expected effect of the chemicals agreed with the stiffness measurements reported by our microchip. Such a low-cost electronic assay that rapidly measures mechanical properties enables previously infeasible studies to advance the science of mechanobiology.

Brain metastases are the most commonly diagnosed type of central nervous system tumor, yet the mechanisms of their occurrence are still widely unknown. Lung cancer, breast cancer, and melanoma are the most common etiologies, but renal and colorectal cancers have also been described as metastasizing to the brain. Regardless of their origin, there are common mechanisms for progression to all types of brain metastases, such as the creation of a suitable tumor microenvironment in the brain, priming of tumor cells, adaptations to survive spreading in lymphatic and blood vessels, and development of mechanisms to penetrate the blood-brain barrier. However, there are complex genetic and molecular interactions that are specific to every type of primary tumor, making the understanding of the metastatic progression of tumors to the brain a challenging field of study. In this review, we aim to summarize current knowledge on the pathophysiology of brain metastases, from specific genetic characteristics of commonly metastatic tumors to the molecular and cellular mechanisms involved in progression to the central nervous system. We also briefly discuss current challenges in targeted therapies for brain metastases and how there is still a gap in knowledge that needs to be overcome to improve patient outcomes.

Tension-adhesion interplay is a crucial mechanism in multicellular organisms that determines the tension differential among internal and external interfaces, which in turn, mediates tissue surface tension and cell sorting, morphogenesis and remodeling, and cancer progression. Cadherins are widely believed to be involved, yet key aspects of the process are neither well characterized nor quantified. This study demonstrates the critical role of N-cadherin in driving tension polarization throughout the actin cortical network. N-cadherin regulates both tension increase at the cell-medium (external) interface and decrease at the cell-cell (internal) interface, and their quantitative magnitudes, both absolute and relative, strongly depend on the surface density of N-cadherin. Furthermore, the strength of tension polarization also increases with respect to the number of cell-cell interfaces for cells within a multicellular cluster. The cadherin-actin contractility linkage is mediated by Rac1, which serves as a molecular switch to trigger cortex remodeling and contraction via myosin II. Inhibition of Rac1 activity decreases tension polarization and leads to reduced coherence in both small clusters and spheroids. These results provide a pathway to reconcile opposing theories for tissue surface tension generation and perspectives in cancer treatment.

The cancer metastatic cascade includes a series of mechanical barrier-crossing events, involving the physical movement of cancer cells from their primary location to a distant organ. This review describes the physical changes that influence tumour proliferation, progression, and metastasis. We identify potential mechanical signatures at every step of the metastatic cascade and discuss some latest mechanobiology-based therapeutic interventions to highlight the importance of interdisciplinary approaches in cancer diagnosis and treatment.

Circulating tumor cells (CTCs) are rare (a few cells per milliliter of blood) and mostly isolated as single-cell CTCs (scCTCs). CTC clusters (cCTCs), even rarer, are of growing interest, notably because of their higher metastatic potential, but very difficult to isolate.

We introduce gravity-based microfiltration (GµF) for facile isolation of cCTCs using in-house fabricated microfilters and 3D printed cartridges. Optimal flow rate and pore size for cCTC isolation are determined by GµF of cultured ovarian single cells and cell clusters spiked in healthy blood. We perform GµF of blood from orthotopic ovarian cancer mouse models and characterize the morphological features of scCTCs and cCTCs, and the expression of molecular markers for aggressiveness. Finally, we analyze blood from 17 epithelial ovarian cancer patients with either localized or metastatic disease, and from 13 colorectal cancer liver metastasis patients.

Here, we show that GµF optimized for cell cluster isolation captures cCTCs from blood while minimizing unwanted cluster disaggregation, with ~85% capture efficiency. We detect cCTCs in every patient, with between 2-100+ cells. We find cCTCs represent between 5-30% of all CTC capture events, and 10-80% of the CTCs are clustered; remarkably, in 10 patients, most CTCs are circulating not as scCTCs, but as cCTCs.

GµF uncovers the unexpected prevalence and frequency of cCTCs including sometimes very large ones in epithelial ovarian cancer patients, and motivates additional studies to uncover their properties and role in disease progression.

Despite being discovered decades ago, metastasis remains a formidable challenge in cancer treatment. During the intermediate phase of metastasis, tumor cells detach from primary tumor or metastatic sites and travel through the bloodstream and lymphatic system to distant tissues. These tumor cells in the circulation are known as circulating tumor cells (CTCs), and a higher number of CTCs has been linked to poor prognoses in various cancers. The blood is an inhospitable environment for any foreign cells, including CTCs, as they face numerous challenges, such as the shear stress within blood vessels and their interactions with blood and immune cells. However, the exact mechanisms by which CTCs survive the hostile conditions of the bloodstream remain enigmatic. Platelets have been studied for their interactions with tumor cells, promoting their survival, growth, and metastasis. This review explores the latest clinical methods for enumerating CTCs, recent findings on platelet-CTC crosstalk, and current research on antiplatelet therapy as a potential strategy to inhibit metastasis, offering new therapeutic insights.

Laboratory and clinical data have provided insights into the role of platelets in promoting CTC survival, while clinical advancements in CTC enumeration offer improved prognostic tools.

CTCs play a critical role in metastasis, and their interactions with platelets aid their survival in the hostile environment of the bloodstream. Understanding this crosstalk offers insights into potential therapeutic strategies, including antiplatelet therapy, to inhibit metastasis and improve cancer treatment outcomes.

Gastric cancer (GC) presents a significant global health burden, with metastasis being the leading cause of treatment failure and mortality. NAT10, a regulatory protein involved in mRNA acetylation, has been implicated in various cancers. However, its role in GC, especially concerning metastasis and immune interactions, remains unclear. Utilizing multi-omics data from gastric cancer samples, we conducted comprehensive analyses to investigate NAT10 expression, its correlation with clinical parameters and immune relevance. Bioinformatics analysis and digital image processing were employed for this purpose. Furthermore, in vitro and in vivo experiments were conducted to elucidate the functional role of NAT10 in gastric cancer progression, aiming to provide deeper biological insights. Our findings reveal a significant association between NAT10 expression and various aspects of transcriptional, protein, as well as tumor immunity in GC patients. Additionally, we demonstrated that NAT10 promotes gastric cancer cell proliferation and migration, both in cellular models and in animal studies, suggesting its involvement in early tumor microvascular metastasis. NAT10 emerges as a promising molecular target, offering potential avenues for further research into molecular mechanisms and therapeutic strategies for GC.

Circulating tumor cells (CTCs) and their clusters are the drivers of metastasis, but we have an incomplete understanding of how they interact with capillary beds. Using microfluidic models mimicking human capillary bifurcations, we observed cell size- and bifurcation-dependent shedding of nuclei-free fragments by patient CTCs, CTC-derived explant cells and numerous cancer cell lines. Shedding reduced cell sizes up to 61%, facilitating their transit through bifurcations. We demonstrated that shed fragments were a novel subclass of large extracellular vesicles (LEVs), "shearosomes", that require shear stress for their biogenesis and whose proteome was associated with immune-related pathways. Shearosomes exhibited functions characteristic of previously identified EVs including cell-directed internalization by endothelial and immune cells, and intercellular communication abilities such as disruption of endothelial barrier integrity, polarization of monocytes into M2 tumor-promoting macrophages and interactions between endothelial and immune cells. Cumulatively, these findings suggest that CTCs shed shearosomes in capillary beds that drive key processes involved in the formation of pre-metastatic niches.

The circulation of tumor cells through the bloodstream is a significant step in tumor metastasis. To better understand the metastatic process, circulating tumor cell (CTC) survival in the circulation must be explored. While immune interactions with CTCs in recent decades have been examined, research has yet to sufficiently explain some CTC behaviors in blood flow. Studies related to CTC mechanical responses in the bloodstream have recently been conducted to further study conditions under which CTCs might die. While experimental methods can assess the mechanical properties and death of CTCs, increasingly sophisticated computational models are being built to simulate the blood flow and CTC mechanical deformation under fluid shear stresses (FSS) in the bloodstream.Several factors contribute to the mechanical deformation and death of CTCs as they circulate. While FSS can damage CTC structure, diverse interactions between CTCs and blood components may either promote or hinder the next metastatic step-extravasation at a remote site. Overall understanding of how these factors influence the deformation and death of CTCs could serve as a basis for future experiments and simulations, enabling researchers to predict CTC death more accurately. Ultimately, these efforts can lead to improved metastasis-specific therapeutics and diagnostics specific in the future.

Lung cancer, the leading cause of death worldwide, is associated with the highest morbidity. Non-small cell lung cancer (NSCLC) accounts for 80-85% of lung cancer cases. Advances in the domain of cancer treatment have improved the prognosis and quality of life of patients with metastatic NSCLC. Nevertheless, tumor progression or metastasis owing to treatment failure caused by primary or secondary drug resistance remains the cause of death in the majority of cases. Epithelial-mesenchymal transition (EMT), a vital biological process wherein epithelial cancer cells lose their inherent adhesion and transform into more invasive mesenchymal-like cells, acts as a powerful engine driving tumor metastasis. EMT can also induce immunosuppression in the tumor environment, thereby promoting cancer development and poor prognosis among patients with NSCLC. This review aims to elucidate the effect of EMT on metastasis and the tumor immune microenvironment. Furthermore, it explores the possible roles of EMT inhibition in improving the treatment efficacy of NSCLC. Targeting EMT may be an ideal mechanism to inhibit tumor growth and progression at multiple steps.

Metastatic disease and cancer recurrence are the primary causes of cancer-related deaths. Circulating tumor cells (CTCs) and disseminated tumor cells (DTCs) are the driving forces behind the spread of cancer cells. The emergence and development of liquid biopsy using rare CTCs as a minimally invasive strategy for early-stage tumor detection and improved tumor management is a promising advancement in recent years. However, before blood sample analysis and clinical translation, precise isolation of CTCs from patients' blood based on their biophysical properties, followed by molecular identification of CTCs using single-cell multi-omics technologies is necessary to understand tumor heterogeneity and provide effective diagnosis and monitoring of cancer progression. Additionally, understanding the origin, morphological variation, and interaction between CTCs and the primary and metastatic tumor niche, as well as and regulatory immune cells, will offer new insights into the development of CTC-based advanced tumor targeting in the future clinical trials.

Breast cancer (BC) bone metastasis poses a significant clinical challenge due to its impact on patient prognosis and quality of life. Curcumin (CUR), a natural polyphenol compound found in turmeric, has shown potential in cancer therapy due to its anti-inflammatory, antioxidant, and anticancer properties. However, its metabolic instability and hydrophobicity have hindered its clinical applications, leading to a short plasma half-life, poor absorption, and low bioavailability. To enhance the drug-like properties of CUR, nanotechnology-based delivery strategies have been employed, utilizing polymeric, lipidic, and inorganic nanoparticles (NPs). These approaches have effectively overcome CUR's inherent limitations by enhancing its stability and cellular bioavailability both in vitro and in vivo. Moreover, targeting molecules with high selectivity towards bone metastasized breast cancer cells can be used for site specific delivery of curcumin. Alendronate (ALN), a bone-seeking bisphosphonate, is one such moiety with high selectivity towards bone and thus can be effectively used for targeted delivery of curcumin loaded nanocarriers. This review will detail the process of bone metastasis in BC, elucidate the mechanism of action of CUR, and assess the efficacy of nanotechnology-based strategies for CUR delivery. Specifically, it will focus on how these strategies enhance CUR's stability and improve targeted delivery approaches in the treatment of BC bone metastasis.

Understanding the metastatic cascade is critical for the treatment and prevention of cancer-related death. Within a tumor, immune cells have the capacity to fuse with tumor cells to generate tumor-immune hybrid cells (THCs). THCs are hypothesized to be a subset of cancer cells with the capacity to enter circulation as circulating hybrid cells (CHC) and seed metastases. To understand the mechanism of THC metastasis, we investigated CHCs in peripheral blood from patients with stage IV colorectal cancer (CRC), as well as THCs in tissues of primary colorectal cancers and their liver metastasis sites using immunofluorescence, spatial proteomic, spatial transcriptomic, molecular classification, and molecular pathway analyses. Our findings indicated a high prevalence of CHCs and THCs in patients with stage IV CRC. THCs expressed CTLA4 in primary CRC lesions and correlated with upregulation of CD68, CD4, and HLA-DR in metastatic liver lesions, which is found in the consensus molecular subtype (CMS) 1 of primary CRC tissue. Pathway analysis of these genes suggested that THCs are associated with neutrophils due to upregulation of neutrophil extracellular trap signaling (NET) and neutrophil degranulation pathways. These data provide molecular pathways for the formation of THCs suggesting fusion with neutrophils, which may facilitate extravasation and metastatic seeding.

Screening liters of blood (i.e., apheresis) represents a generalized approach to promote the reliable access to circulating tumor cell clusters (CTCCs), which are known to be highly metastasis-competent, yet ultrarare. However, no existing CTCC sorting technology has demonstrated high throughput, high yield, low shear stress, and minimal blood dilution simultaneously as required in apheresis. Here, a label-free method is introduced termed Precision Apheresis for Non-invasive Debulking of cell Aggregates (PANDA) to continuously isolate CTCCs from undiluted blood to clean buffer through size sorting, processing 1.4 billion cells per second. The cell focusing is optimized within whole blood leveraging secondary transverse flow and margination. The PANDA chip recovers >90% of spiked ≈24 rare HeLa cell clusters from 100 mL undiluted blood samples (equivalent to ≈500 billion blood cells) at 1 L h-1 throughput, with ≤20s device residence time, ≤15 Pa shear stress, and >99.9% return of blood components. The technology lays the groundwork for future routine isolation to increase the recovery of these ultrarare yet clinically significant tumor cell populations from large volumes of blood to advance cancer research, early detection, and treatment.

Bone metastasis is the prevalent form of metastasis in breast cancer, resulting in severe pain, pathological fractures, nerve compression, hypercalcemia, and other complications that significantly impair patients' quality of life. The infiltration and colonization of breast cancer (BC) cells in bone tissue disrupt the delicate balance between osteoblasts and osteoclasts within the bone microenvironment, initiating a vicious cycle of bone metastasis. Once bone metastasis occurs, conventional medical therapy with bone-modifying agents is commonly used to alleviate bone-related complications and improve patients' quality of life. However, the utilization of bone-modifying agents may cause severe drug-related adverse effects. Plant-derived natural products such as terpenoids, alkaloids, coumarins, and phenols have anti-tumor, anti-inflammatory, and anti-angiogenic pharmacological properties with minimal side effects. Certain natural products that exhibit both anti-breast cancer and anti-bone metastasis effects are potential therapeutic agents for breast cancer bone metastasis (BCBM). This article reviewed the effects of plant-derived natural products against BCBM and their mechanisms to provide a reference for the research and development of drugs related to BCBM.

Prostate cancer is a disease in which cells in the prostate, a gland in the male reproductive system below the bladder, grow out of control and, among men, it is the second-most frequently diagnosed cancer (other than skin cancer). In recent years, prostate cancer death rate has stabilized and, currently, it is the second-most frequent cause of cancer death in men (after lung cancer). Most deaths occur due to metastasis, as cancer cells from the original tumor establish secondary tumors in distant organs. For a long time, classical cell cultures and animal models have been utilized in basic and applied scientific research, including clinical applications for many diseases, such as prostate cancer, since no better alternatives were available. Although helpful in dissecting cellular mechanisms, these models are poor predictors of physiological behavior mainly because of the lack of appropriate microenvironments. Microfluidics has emerged in the last two decades as a technology that could lead to a paradigm shift in life sciences and, in particular, controlling cancer. Microfluidic systems, such as organ-on-chips, have been assembled to mimic the critical functions of human organs. These microphysiological systems enable the long-term maintenance of cellular co-cultures in vitro to reconstitute in vivo tissue-level microenvironments, bridging the gap between traditional cell cultures and animal models. Several reviews on microfluidics for prostate cancer studies have been published focusing on technology advancement and disease progression. As metastatic castration-resistant prostate cancer remains a clinically challenging late-stage cancer, with no curative treatments, we expanded this review to cover recent microfluidic applications related to prostate cancer research. The review includes discussions of the roles of microfluidics in modeling the human prostate, prostate cancer initiation and development, as well as prostate cancer detection and therapy, highlighting potentially major contributions of microfluidics in the continuous march toward eradicating prostate cancer.

Liver metastasis remains the primary cause of mortality in patients with colon cancer. Identifying specific driver gene mutations that contribute to metastasis may offer viable therapeutic targets. To explore clonal evolution and genetic heterogeneity within the metastasis, we conducted single-cell exome sequencing on 150 single cells isolated from the primary tumor, liver metastasis, and lymphatic metastasis from a stage IV colon cancer patient. The genetic landscape of the tumor samples revealed that both lymphatic and liver metastases originated from the same region of the primary tumor. Notably, the liver metastasis was derived directly from the primary tumor, bypassing the lymph nodes. Comparative analysis of the sequencing data for individual cell pairs within different tumors demonstrated that the genetic heterogeneity of both liver and lymphatic metastases was also greater than that of the primary tumor. This finding indicates that liver and lymphatic metastases arose from clusters of circulating tumor cell (CTC) of a polyclonal origin, rather than from a single cell from the primary tumor. Single-cell transcriptome analysis suggested that higher EMT score and CNV scores were associated with more polyclonal metastasis. Additionally, a mutation in the TRPS1 (Transcriptional repressor GATA binding 1) gene, TRPS1 R544Q, was enriched in the single cells from the liver metastasis. The mutation significantly increased CRC invasion and migration both in vitro and in vivo through the TRPS1R544Q/ZEB1 axis. Further TRPS1 mutations were detected in additional colon cancer cases, correlating with advanced-stage disease and inferior prognosis. These results reveal polyclonal seeding and TRPS1 mutation as potential mechanisms driving the development of liver metastases in colon cancer.

Circulating tumor cells (CTCs) currently represent a revolutionary tool offering unique insights for the evaluation of cancer progression, metastasis, and response to therapies. Indeed, CTCs, upon detachment from primary tumors, enter the bloodstream and acquire a great potential for their use for personalized cancer management. In this review, we describe the current understanding of and advances in the clinical employment of CTCs. Although considered rare and fleeting, CTCs are now recognized as key players favoring the development of cancer metastasis and disease recurrence, particularly in malignant melanoma, lung, breast, and colorectal cancer patients. To date, the advancements in technology and the development of several successful approaches, also including immunomagnetic enrichment allow for a reliable and reproducible detection and characterization of CTCs. Those innovative methodologies improved the isolation, quantification, and characterization of CTCs from the blood of cancer patients, providing extremely useful evidence and new insights into the nature of the tumor, its epithelial/mesenchymal profile, and its potential resistance to therapy. In fact, in addition to their prognostic and predictive value, CTCs could serve as a valuable instrument for real-time monitoring of treatment response and disease recurrence, facilitating timely interventions and thus improving patient outcomes. However, despite their potential, several challenges hinder the widespread clinical utility of CTCs: (i) CTCs' rarity and heterogeneity pose technical limitations in isolation and characterization, as well as significant hurdles in their clinical implementation; (ii) it is mandatory to standardize CTC detection methods, optimize the sample processing techniques, and integrate them with existing diagnostic modalities; and (iii) the need for the development of new techniques, such as single-cell analysis platforms, to enhance the sensitivity and specificity of CTC detection, thereby facilitating their integration into routine clinical practice. In conclusion, CTCs represent a potential extraordinary tool in cancer diagnostics and therapeutics, offering unprecedented opportunities for personalized medicine and precision oncology. Moreover, their ability to provide real-time insights into tumor biology, treatment response, and disease progression underlines a great potential for their clinical application to improve patients' outcomes and advance our understanding of cancer biology.

Circulating tumour cells (CTCs) and CTC clusters are considered metastatic precursors due to their ability to seed distant metastasis. However, navigating the bloodstream presents a significant challenge for CTCs, as they must endure fluid shear forces and resist detachment-induced anoikis. Consequently, while a large number of cells from the primary tumour may enter the circulation, only a tiny fraction will result in metastasis. Nevertheless, the metastatic potency dramatically increases when CTCs travel in conjunction with other cell types to form CTC clusters, including neutrophils, myeloid-derived suppressor cells, macrophages, platelets, cancer-associated fibroblasts and red blood cells found in circulation. Such heterotypic CTC clustering events have been identified in a variety of cancer types and may serve as intriguing therapeutic targets and novel biomarkers for liquid biopsy. This review summarises recent advances in microfluidic technologies designed for the isolation of CTC clusters and explores the biological properties of distinct types of CTC clusters within the circulatory system. Investigation of the mechanisms of CTC cluster-blood microenvironment interactions may offer a promising avenue for gaining fresh insights into CTC cluster-mediated metastatic progression and reveal potential opportunities for devising personalised antimetastasis treatments.

The primary reason for high mortality rates among cancer patients is metastasis, where tumor cells migrate through the bloodstream from the original site to other parts of the body. Recent advancements in technology have significantly enhanced our comprehension of the mechanisms behind the bloodborne spread of circulating tumor cells (CTCs). One critical process, DNA methylation, regulates gene expression and chromosome stability, thus maintaining dynamic equilibrium in the body. Global hypomethylation and locus-specific hypermethylation are examples of changes in DNA methylation patterns that are pivotal to carcinogenesis. This comprehensive review first provides an overview of the various processes that contribute to the formation of CTCs, including epithelial-mesenchymal transition (EMT), immune surveillance, and colonization. We then conduct an in-depth analysis of how modifications in DNA methylation within CTCs impact each of these critical stages during CTC dissemination. Furthermore, we explored potential clinical implications of changes in DNA methylation in CTCs for patients with cancer. By understanding these epigenetic modifications, we can gain insights into the metastatic process and identify new biomarkers for early detection, prognosis, and targeted therapies. This review aims to bridge the gap between basic research and clinical application, highlighting the significance of DNA methylation in the context of cancer metastasis and offering new avenues for improving patient outcomes.

Many tumors prefer to metastasize to bone, but the underlying mechanisms remain elusive. The human skeletal system has unique physical properties, that are distinct from other organs, which play a key role in directing the behavior of tumor cells within bone. Understanding the physical journey of tumor cells within bone is crucial. In this review we discuss bone metastasis in the context of how physical cues in the bone vasculature and bone marrow niche regulate the fate of tumor cells. Our objective is to inspire innovative diagnostic and therapeutic approaches for bone metastasis from a mechanobiological perspective.

Metastasis remains the principal trigger for relapse and mortality across diverse cancer types. Circulating tumor cells (CTCs), which originate from the primary tumor or its metastatic sites, traverse the vascular system, serving as precursors in cancer recurrence and metastasis. Nevertheless, before CTCs can establish themselves in the distant parenchyma, they must overcome significant challenges present within the circulatory system, including hydrodynamic shear stress (HSS), oxidative damage, anoikis, and immune surveillance. Recently, there has been a growing body of compelling evidence suggesting that a specific subset of CTCs can persist within the bloodstream, but the precise mechanisms of their survival remain largely elusive. This review aims to present an outline of the survival challenges encountered by CTCs and to summarize the recent advancements in understanding the underlying survival mechanisms, suggesting their implications for cancer treatment.

Most cancer-related deaths are due to the spread of tumour cells throughout the body-a process known as metastasis. While in the vasculature, these cells are referred to as circulating tumour cells (CTCs) and can be found as either single cells or clusters of cells (often including platelets), with the latter having the highest metastatic potential. However, the biology of CTC clusters is poorly understood, and there are no therapies that specifically target them. We previously developed an in vitro model system for CTC clusters and proposed a new extravasation model that involves cluster dissociation, adherence, and single-cell invasion in response to TGF-β1 released by platelets. Here, we investigated TGF-β1-induced gene expression changes in this model, focusing on genes for which targeted drugs are available. In addition to the upregulation of the TGF-β1 signalling pathway, we found that (i) genes in the EGF/EGFR pathway, including those coding for EGFR and several EGFR ligands, were also induced, and (ii) Erlotinib and Osimertinib, two therapeutic EGFR/tyrosine kinase inhibitors, decreased the TGF-β1-induced adherence and invasion of the CTC cluster-like line despite the line expressing wild-type EGFR. Overall, we suggest that EGFR inhibitors have the potential to decrease the dispersal of CTC clusters that respond to TGF-β1 and overexpress EGFR (irrespective of its status) and thus could improve patient survival.

During the metastatic cascade, cancer cells travel through the bloodstream as circulating tumor cells (CTCs) to a secondary site. Clustered CTCs have greater shear stress and treatment resistance, yet their biology remains poorly understood. We therefore engineered a tunable superhydrophobic array device (SHArD). The SHArD-C was applied to culture a clinically relevant model of CTC clusters. Using our device, we cultured a model of cancer cell aggregates of various sizes with immortalized cancer cell lines. These exhibited higher E-cadherin expression and are significantly more capable of surviving high fluid shear stress-related forces compared to single cells and model clusters grown using the control method, helping to explain why clustering may provide a metastatic advantage. Additionally, the SHArD-S, when compared with the AggreWell 800 method, provides a more consistent spheroid-forming device culturing reproducible sizes of spheroids for multiple cancer cell lines. Overall, we designed, fabricated, and validated an easily tunable engineered device which grows physiologically relevant three-dimensional (3D) cancer models containing tens to thousands of cells.

Early evaluation of tumor heterogeneity related to metastasis and outcomes is a major challenge in the management of advanced breast cancer (BCa) in the clinic. In this study, we introduced the value of baseline circulating tumor cells (CTC) and ctDNA for early differentiation of clinical stages, tumor heterogeneity, and prognosis in clinic.

A total of 292 patients with BCa were enrolled in this study, including 254 Stage IV and 38 Stage III patients, and examined the baseline levels of CTCs, CTC-clusters, and plasma ctDNA before initiating therapies. Outcomes including progression-free survival (PFS) and overall survival were evaluated using proportional hazards regression analysis.

The baseline CTCs, including HER2+ CTCs, in Stage IV patients were approximately 9.5 times higher than those detected in Stage III patients. Baseline CTC counts with a cutoff of 5 were significantly associated with the prognosis. Within each stage, patients with <5 CTCs had significantly longer PFS. Stage III patients with no CTCs exhibited the longest survival compared with patients with ≥1 CTC. CTC-clusters were only found in Stage IV patients, among whom 15 Stage IV patients with ≥5 CTC-clusters had the worst PFS compared with the 239 Stage IV patients with <5 CTC-clusters. Similar outcomes were observed in 28 out of 254 Stage IV patients who had at least one CTC-cluster detected, as these patients had shorter PFS compared with CTC-cluster negative group. The major differences in ctDNA mutations between patients with Stage III and Stage IV BCa were in PIK3CA and ESR1, which were associated with specific organ metastasis and worse outcomes.

Assessing the baseline levels of CTCs, CTC-clusters, and mutational ctDNA profile could reliably aid in differentiation of clinical stage and early prediction of metastasis and outcomes in advanced BCa.

The metastatic cascade includes a blood circulation step for cells detached from the primary tumor. This stage involves significant shear stress as well as large and fast deformation as the cells circulate through the microvasculature. These mechanical stimuli are well reproduced in microfluidic devices. However, the recovery dynamics after deformation is also pivotal to understand how a cell can pass through the multiple capillary constrictions encountered during a single hemodynamic cycle. The microfluidic system developed in this work allows single cell recovery to be studied under flow-free conditions following pressure-actuated cell deformation inside constricted microchannels. We used three breast cancer cell lines - namely MCF-7, SK-BR3 and MDA-MB231 - as cellular models representative of different cancer phenotypes. Changing the size of the constriction allows exploration of moderate to strong deformation regimes, the latter being associated with the formation of plasma membrane blebs. In the regime of moderate deformation, all cell types display a fast elastic recovery behavior followed by a slower viscoelastic regime, well described by a double exponential decay. Among the three cell types, cells of the mesenchymal phenotype, i.e. the MDA-MB231 cells, are softer and the most fluid-like, in agreement with previous studies. Our main finding here is that the fast elastic recovery regime revealed by our novel microfluidic system is under the control of cell contractility ensured by the integrity of the cell cortex. Our results suggest that the cell cortex plays a major role in the transit of circulating tumor cells by allowing their fast morphological recovery after deformation in blood capillaries.

Microfluidic chips are valuable tools for studying intricate cellular and cell-microenvironment interactions. Traditional in vitro cancer models lack accuracy in mimicking the complexities of in vivo tumor microenvironment. However, cancer-metastasis-on-a-chip (CMoC) models combine the advantages of 3D cultures and microfluidic technology, serving as powerful platforms for exploring cancer mechanisms and facilitating drug screening. These chips are able to compartmentalize the metastatic cascade, deepening the understanding of its underlying mechanisms. This article provides an overview of current CMoC models, focusing on distinctive models that simulate invasion, intravasation, circulation, extravasation, and colonization, and their applications in drug screening. Furthermore, challenges faced by CMoC and microfluidic technologies are discussed, while exploring promising future directions in cancer research. The ongoing development and integration of these models into cancer studies are expected to drive transformative advancements in the field.

The characterization of circulating tumor cells (CTC) and circulating tumor microemboli (CTM) has emerged as both a challenge to the standard view of metastasis, and as a valuable means for understanding genotypic and phenotypic variability shown even within the same cancer type. However, in the case of salivary gland neoplasms, limited data are available for the role that CTCs and CTMs play in metastasis and secondary tumor formation.ru.AQ1 In response to this, we propose that similarities between in vitro clusters of cultured salivary gland cancer cells may act as a surrogate model for in vivo CTCs and CTMs isolated from patients.

Using techniques in immunofluorescence, immunoblotting, and 2-dimensional migration, we isolated and characterized a group of cohort cells from a commercially available cell line (HTB-41). Results: Here, cells exhibited a hybrid phenotype with simultaneous expression of both epithelial and mesenchymal markers (E-cadherin, vimentin, and α-SMA). Cohort cells also exhibited increased migration in comparison to parental cells.

Data suggest that these isolated cell clusters may fucntion as a potential in vitro model of CTCs and CTMs.

Metastasis remains the leading cause of cancer deaths worldwide and lung cancer, known for its highly metastatic progression, remains among the most lethal of malignancies. Lung cancer metastasis can selectively spread to multiple different organs, however the genetic and molecular drivers for this process are still poorly understood. Understanding the heterogeneous genomic profile of lung cancer metastases is considered key in identifying therapeutic targets that prevent its spread. Research has identified the key source for metastasis being clusters of cells rather than individual cancer cells. These clusters, known as metastatic cancer cell clusters (MCCCs) have been shown to be 100-fold more tumorigenic than individual cancer cells. Unfortunately, access to these primary drivers of metastases remains difficult and has limited our understanding of their molecular and genomic profiles. Strong evidence in the literature suggests that differentially regulated biological pathways in MCCCs can provide new therapeutic drug targets to help combat cancer metastases. In order to expand research into MCCCs and their role in metastasis, we demonstrate a novel, proof of principle technology, to capture MCCCs directly from patients' whole blood. Our platform can be readily tuned for different solid tumor types by combining a biomimicry-based margination effect coupled with immunoaffinity to isolate MCCCs. Adopting a selective capture approach based on overexpressed CD44 in MCCCs provides a methodology that preferentially isolates them from whole blood. Furthermore, we demonstrate a high capture efficiency of more than 90% when spiking MCCC-like model cell clusters into whole blood. Characterization of the captured MCCCs from lung cancer patients by immunofluorescence staining and genomic analyses, suggests highly differential morphologies and genomic profiles. This study lays the foundation to identify potential drug targets thus unlocking a new area of anti-metastatic therapeutics.

During metastatic dissemination, circulating tumour cells (CTCs) enter capillary beds, where they experience mechanical constriction forces. The transient and persistent effects of these forces on CTCs behaviour remain poorly understood. Here, we developed a high-throughput microfluidic platform mimicking human capillaries to investigate the impact of mechanical constriction forces on malignant and normal breast cell lines. We observed that capillary constrictions induced nuclear envelope rupture in both cancer and normal cells, leading to transient changes in nuclear and cytoplasmic area. Constriction forces transiently activated cGAS/STING and pathways involved in inflammation (NF-κB, STAT and IRF3), especially in the non-malignant cell line. Furthermore, the non-malignant cell line experienced transcriptional changes, particularly downregulation of epithelial markers, while the metastatic cell lines showed minimal alterations. These findings suggest that mechanical constriction forces within capillaries may promote differential effects in malignant and normal cell lines.

Globally, lung cancer stands as the leading type of cancer in terms of incidence and is the major source of mortality attributed to cancer. We have outlined the molecular biomarkers for lung cancer that are available clinically. Circulating tumor cells (CTCs) spread from the original location, circulate in the bloodstream, extravasate, and metastasize, forming secondary tumors by invading and establishing a favorable environment. CTC analysis is considered a common liquid biopsy method for lung cancer. We have enumerated both in vivo and ex vivo techniques for CTC separation and enrichment, examined the advantages and limitations of these methods, and also discussed the detection of CTCs in other bodily fluids. We have evaluated the value of CTCs, as well as CTCs in conjunction with other biomarkers, for their utility in the early detection and prognostic assessment of patients with lung cancer. CTCs engage with diverse cells of the metastatic process, interfering with the interaction between CTCs and various cells in metastasis, potentially halting metastasis and enhancing patient prognosis.

After reading the review by An et al "Biological factors driving colorectal cancer metastasis", which covers the problem of the metastasis of colorectal cancer (CRC), I had a desire to discuss with readers one of the exciting problems associated with dormant metastases. Most deaths from CRCs are caused by metastases, which can be detected both at diagnosis of the primary tumor and several years or even decades after treatment. This is because tumor cells that enter the bloodstream can be destroyed by the immune system, cause metastatic growth, or remain dormant for a long time. Dormant tumor cells may not manifest themselves throughout a person's life or, after some time and under appropriate conditions, may give rise to the growth of metastases. In this editorial, we will discuss the most important features of dormant metastases and the mechanisms of premetastatic niche formation, as well as factors that contribute to the activation of dormant metastases in CRCs. We will pay special attention to the possible mechanisms involved in the formation of circulating tumor cell complexes and the choice of therapeutic strategies that promote the dormancy or destruction of tumor cells in CRCs.

During cancer metastasis, cancer cells will encounter various microenvironments with diverse physical characteristics. Changes in these physical characteristics such as tension, stiffness, viscosity, compression, and fluid shear can generate biomechanical cues that affect cancer cells, dynamically influencing numerous pathophysiological mechanisms. For example, a dense extracellular matrix drives cancer cells to reorganize their cytoskeleton structures, facilitating confined migration, while this dense and restricted space also acts as a physical barrier that potentially results in nuclear rupture. Identifying these pathophysiological processes and understanding their underlying mechanobiological mechanisms can aid in the development of more effective therapeutics targeted to cancer metastasis. In this review, we outline the advances of engineering microfluidic devices in vitro and their role in replicating tumor microenvironment to mimic in vivo settings. We highlight the potential cellular mechanisms that mediate their ability to adapt to different microenvironments. Meanwhile, we also discuss some important mechanical cues that still remain challenging to replicate in current microfluidic devices in future direction. While much remains to be explored about cancer mechanobiology, we believe the developments of microfluidic devices will reveal how these physical cues impact the behaviors of cancer cells. It will be crucial in the understanding of cancer metastasis, and potentially contributing to better drug development and cancer therapy.

Circulating tumor cell (CTC) clusters play a critical role in carcinoma metastasis. However, the rarity of CTC clusters and the limitations of capture techniques have retarded the research progress. In vitro CTC clusters model can help to further understand the biological properties of CTC clusters and their clinical significance. Therefore, it is necessary to establish reliable in vitro methodological models to form CTC clusters whose biological characteristics are very similar to clinical CTC clusters.

The assays of immunofluorescence, transmission electron microscopy, EdU incorporation, cell adhension and microfluidic chips were used. The experimental metastasis model in mice was used.

We systematically optimized the culture methods to form in vitro CTC clusters model, and more importantly, evaluated it with reference to the biological capabilities of reported clinical CTC clusters. In vitro CTC clusters exhibited a high degree of similarity to the reported pathological characteristics of CTC clusters isolated from patients at different stages of tumor metastasis, including the appearance morphology, size, adhesive and tight junctions-associated proteins, and other indicators of CTC clusters. Furthermore, in vivo experiments also demonstrated that the CTC clusters had an enhanced ability to grow and metastasize compared to single CTC.

The study provides a reliable model to help to obtain comparatively stable and qualified CTC clusters in vitro, propelling the studies on tumor metastasis.

Metastatic tumors have poor prognoses for progression-free and overall survival for all cancer patients. Rare circulating tumor cells (CTCs) and rarer circulating tumor cell clusters (CTCCs) are potential biomarkers of metastatic growth, with CTCCs representing an increased risk factor for metastasis. Current detection platforms are optimized for ex vivo detection of CTCs only. Microfluidic chips and size exclusion methods have been proposed for CTCC detection; however, they lack in vivo utility and real-time monitoring capability. Confocal backscatter and fluorescence flow cytometry (BSFC) has been used for label-free detection of CTCCs in whole blood based on machine learning (ML) enabled peak classification. Here, we expand to a deep-learning (DL)-based, peak detection and classification model to detect CTCCs in whole blood data. We demonstrate that DL-based BSFC has a low false alarm rate of 0.78 events per min with a high Pearson correlation coefficient of 0.943 between detected events and expected events. DL-based BSFC of whole blood maintains a detection purity of 72% and a sensitivity of 35.3% for both homotypic and heterotypic CTCCs starting at a minimum size of two cells. We also demonstrate through artificial spiking studies that DL-based BSFC is sensitive to changes in the number of CTCCs present in the samples and does not add variability in detection beyond the expected variability from Poisson statistics. The performance established by DL-based BSFC motivates its use for in vivo detection of CTCCs. Using transfer learning, we additionally validate DL-based BSFC on blood samples from different species and cancer cell types. Further developments of label-free BSFC to enhance throughput could lead to critical applications in the clinical detection of CTCCs and ex vivo isolation of CTCC from whole blood with minimal disruption and processing steps.

Breast cancer is a significant and deadly threat to women globally. Moreover, Breast cancer metastasis is a complicated process involving multiple biological stages, which is considered a substantial cause of death, where cancer cells spread from the original tumor to other organs in the body-representing the primary mortality factor. Circulating tumor cells (CTCs) are cancer cells detached from the primary or metastatic tumor and enter the bloodstream, allowing them to establish new metastatic sites. CTCs can travel alone or in groups called CTC clusters. Studies have shown that CTC clusters have more potential for metastasis and a poorer prognosis than individual CTCs in breast cancer patients. However, our understanding of CTC clusters' formation, structure, function, and detection is still limited. This review summarizes the current knowledge of CTC clusters' biological properties, isolation, and prognostic significance in breast cancer. It also highlights the challenges and future directions for research and clinical application of CTC clusters.

Oral Squamous Cell Carcinoma (OSCC), the sixth most widespread malignancy in the world, accounts for 90% of all cases of oral cancer. The primary risk factors are tobacco chewing, alcohol consumption, viral infection, and genetic modifications. OSCC has a high morbidity rate due to the lack of early diagnostic methods. Nowadays, liquid biopsy plays a vital role in the initial diagnosis of oral cancer. ctNAs extracted from saliva and serum/plasma offer meaningful insights into tumor genetics and dynamics. The interplay of these elements in saliva and serum/plasma showcases their significance in advancing noninvasive, effective OSCC detection and monitoring.

This review mainly focused on the role of liquid biopsy as an emerging point in the diagnosis and prognosis of OSCC and the current advancements and challenges associated with liquid biopsy.

Liquid biopsy is regarded as a new, minimally invasive, real-time monitoring tool for cancer diagnosis and prognosis. Many biomolecules found in bodily fluids, including ctDNA, ctRNA, CTCs, and EVs, are significant biomarkers to identify cancer in its early stages. Despite these groundbreaking strides, challenges persist. Standardization of sample collection, isolation, processing, and detection methods is imperative for ensuring result reproducibility across diverse studies.

Cancer remains a leading cause of mortality worldwide, with metastasis significantly contributing to its lethality. The metastatic spread of tumor cells, primarily through the bloodstream, underscores the importance of circulating tumor cells (CTCs) in oncological research. As a critical component of liquid biopsies, CTCs offer a non-invasive and dynamic window into tumor biology, providing invaluable insights into cancer dissemination, disease progression, and response to treatment. This review article delves into the recent advancements in CTC research, highlighting their emerging role as a biomarker in various cancer types. We explore the latest technologies and methods for CTC isolation and detection, alongside novel approaches to characterizing their biology through genomics, transcriptomics, proteomics, and epigenetic profiling. Additionally, we examine the clinical implementation of these findings, assessing how CTCs are transforming the landscape of cancer diagnosis, prognosis, and management. By offering a comprehensive overview of current developments and potential future directions, this review underscores the significance of CTCs in enhancing our understanding of cancer and in shaping personalized therapeutic strategies, particularly for patients with metastatic disease.