Extracellular Matrix Stiffness Research Articles

Matrix stiffening facilitates stemness of liver cancer stem cells by YAP activation and BMF inhibition

Matrix stiffening is one of the major risk factors for hepatocellular carcinoma (HCC) and drives tumor progression. The extracellular matrix (ECM) stiffness of HCC displays mechanical heterogeneity, with stiffness increasing from the core to the invasive frontier. The distribution of liver cancer stem cells (CSCs) is related to this mechanical property. However, it is not sufficiently understood how heterogeneous matrix stiffness regulates the stemness of CSCs. In this study, we developed an adjustable gelatin/alginate hydrogel to investigate the effect of various matrix stiffnesses on CSC stemness under three-dimensional culture conditions. Gelatin/alginate hydrogel with the stiffness of soft (5 kPa), medium (16 kPa), and stiff (81 kPa) were prepared by altering the concentration of calcium ions. It was found that a stiffer matrix promoted stemness-associated gene expression, reduced drug sensitivity, enhanced sphere-forming and clonogenic ability, and tumorigenic potential. Mechanistically, matrix stiffening facilitates CSC stemness by increasing Yes-associated protein (YAP) activity and inhibiting Bcl-2 modifying factor (BMF) expression. Knockdown of YAP or overexpression of BMF significantly attenuated matrix stiffening-induced stemness, suggesting the involvement of YAP and BMF in this process. Together, our results unravel the regulatory mechanism of heterogeneous matrix stiffness on CSC stemness and also provide a novel therapeutic strategy for eradicating CSCs and improving the efficiency of HCC treatment.

Piezo1 Activation Drives Enhanced Collagen Synthesis in Aged Animal Skin Induced by Poly L-Lactic Acid Fillers.

Poly L-lactic acid (PLLA) fillers stimulate collagen synthesis by activating various immune cells and fibroblasts. Piezo1, an ion channel, responds to mechanical stimuli, including changes in extracellular matrix stiffness, by mediating Ca2+ influx. Given that elevated intracellular Ca2+ levels trigger signaling pathways associated with fibroblast proliferation, Piezo1 is a pivotal regulator of collagen synthesis and tissue fibrosis. The aim of the present study was to investigate the impact of PLLA on dermal collagen synthesis by activating Piezo1 in both an H2O2-induced cellular senescence model in vitro and aged animal skin in vivo. PLLA elevated intracellular Ca2+ levels in senescent fibroblasts, which was attenuated by the Piezo1 inhibitor GsMTx4. Furthermore, PLLA treatment increased the expression of phosphorylated ERK1/2 to total ERK1/2 (pERK1/2/ERK1/2) and phosphorylated AKT to total AKT (pAKT/AKT), indicating enhanced pathway activation. This was accompanied by upregulation of cell cycle-regulating proteins (CDK4 and cyclin D1), promoting the proliferation of senescent fibroblasts. Additionally, PLLA promoted the expression of phosphorylated mTOR/S6K1/4EBP1, TGF-β, and Collagen I/III in senescent fibroblasts, with GsMTx4 treatment mitigating these effects. In aged skin, PLLA treatment similarly upregulated the expression of pERK1/2/ERK1/2, pAKT/AKT, CDK4, cyclin D1, mTOR/S6K1/4EBP1, TGF-β, and Collagen I/III. In summary, our findings suggest Piezo1's involvement in PLLA-induced collagen synthesis, mediated by heightened activation of cell proliferation signaling pathways such as pERK1/2/ERK1/2, pAKT/AKT, and phosphorylated mTOR/S6K1/4EBP1, underscoring the therapeutic potential of PLLA in tissue regeneration.

Mitochondrial activity related genes of mast cells identify poor prognosis and metastasis of ovarian cancer

The pro-tumorigenic or anti-tumorigenic role of tumor infiltrating mast cells (TIMs) in tumors depends not only on the type of cancer and the degree of tumor progression, but also on their location in the tumor bulk. In our investigation, we employed immunohistochemistry to reveal that the mast cells (MCs) in the tumor stroma are positively correlated with metastasis of ovarian cancer (OC), but not in the tumor parenchyma. To delve deeper into the influence of different culture matrix stiffness on MCs’ biological functions within the tumor parenchymal and stromal regions, we conducted a transcriptome analysis of the mouse MC line (P815) cultured in two-dimensional (2D) or three-dimensional (3D) culture system. Further research has found that the softer 3D extracellular matrix stiffness could improve the mitochondrial activity of MCs to promote proliferation by increasing the expression levels of mitochondrial activity-related genes, namely Pet100, atp5md, and Cox7a2. Furthermore, employing LASSO regression analysis, we identified that Pet100 and Cox7a2 were closely associated with the prognosis of OC patients. These two genes were subsequently employed to construct a risk score model, which revealed that the high-risk group model as one of the prognostic factors for OC patients. Additionally, the XCell algorithm analysis showed that the high-risk group displayed a broader spectrum of immune cell infiltrations. Our research revealed that TIMs in the tumor stroma could promote the metastasis of OC, and mitochondrial activity-related proteins Pet100/Cox7a2 can serve as biomarkers for prognostic evaluation of OC.

Hyperforin improves matrix stiffness induced nucleus pulposus inflammatory degeneration by activating mitochondrial fission

ObjectiveThe continuously increasing extracellular matrix stiffness during intervertebral disc degeneration promotes disease progression. In an attempt to obtain novel treatment methods, this study aims to investigate the changes in nucleus pulposus cells under the stimulation of a stiff microenvironment. DesignRNA sequencing and metabolomics experiments were combined to evaluate the primary nucleus pulposus and screen key targets under mechanical biological stimulation. Additionally, small molecules work in vitro were used to confirm the target regulatory effect and investigate the mechanism. In vivo, treatment effects were validated using a rat caudal vertebrae compression model. ResultsOur research results revealed that by activating TRPC6, hyperforin, a herbaceous extract can rescue the inflammatory phenotype caused by the stiff microenvironment, hence reducing intervertebral disc degeneration (IDD). Mechanically, it activates mitochondrial fission to inhibit PFKFB3. ConclusionIn summary, this study reveals the important bridging role of TRPC6 between mechanical stiffness, metabolism, and inflammation in the context of nucleus pulposus degeneration. TRPC6 activation with hyperforin may become a promising treatment for IDD.

The Role of Biophysical Factors in Organ Development: Insights from Current Organoid Models.

Biophysical factors play a fundamental role in human embryonic development. Traditional in vitro models of organogenesis focused on the biochemical environment and did not consider the effects of mechanical forces on developing tissue. While most human tissue has a Young's modulus in the low kilopascal range, the standard cell culture substrate, plasma-treated polystyrene, has a Young's modulus of 3 gigapascals, making it 10,000-100,000 times stiffer than native tissues. Modern in vitro approaches attempt to recapitulate the biophysical niche of native organs and have yielded more clinically relevant models of human tissues. Since Clevers' conception of intestinal organoids in 2009, the field has expanded rapidly, generating stem-cell derived structures, which are transcriptionally similar to fetal tissues, for nearly every organ system in the human body. For this reason, we conjecture that organoids will make their first clinical impact in fetal regenerative medicine as the structures generated ex vivo will better match native fetal tissues. Moreover, autologously sourced transplanted tissues would be able to grow with the developing embryo in a dynamic, fetal environment. As organoid technologies evolve, the resultant tissues will approach the structure and function of adult human organs and may help bridge the gap between preclinical drug candidates and clinically approved therapeutics. In this review, we discuss roles of tissue stiffness, viscoelasticity, and shear forces in organ formation and disease development, suggesting that these physical parameters should be further integrated into organoid models to improve their physiological relevance and therapeutic applicability. It also points to the mechanotransductive Hippo-YAP/TAZ signaling pathway as a key player in the interplay between extracellular matrix stiffness, cellular mechanics, and biochemical pathways. We conclude by highlighting how frontiers in physics can be applied to biology, for example, how quantum entanglement may be applied to better predict spontaneous DNA mutations. In the future, contemporary physical theories may be leveraged to better understand seemingly stochastic events during organogenesis.

Biomimetic Hydrogel-Mediated Mechano-Immunometabolic Therapy for Inhibition of ccRCC Recurrence After Surgery.

The unique physical tumor microenvironment (TME) and aberrant immune metabolic status are two obstacles that must be overcome in cancer immunotherapy to improve clinical outcomes. Here, an in situ mechano-immunometabolic therapy involving the injection of a biomimetic hydrogel is presented with sequential release of the anti-fibrotic agent pirfenidone, which softens the stiff extracellular matrix, and small interfering RNA IDO1, which disrupts kynurenine-mediated immunosuppressive metabolic pathways, together with the multi-kinase inhibitor sorafenib, which induces immunogenic cell death. This combination synergistically augmented tumor immunogenicity and induced anti-tumor immunity. In mouse models of clear cell renal cell carcinoma, a single-dose peritumoral injection of a biomimetic hydrogel facilitated the perioperative TME toward a more immunostimulatory landscape, which prevented tumor relapse post-surgery and prolonged mouse survival. Additionally, the systemic anti-tumor surveillance effect induced by local treatment decreased lung metastasis by inhibiting epithelial-mesenchymal transition conversion. The versatile localized mechano-immunometabolic therapy can serve as a universal strategy for conferring efficient tumoricidal immunity in "cold" tumor postoperative interventions.

Combined effects of matrix stiffness and obesity-associated signaling directs progressive phenotype in PANC-1 pancreatic cancer cells in vitro.

Obesity is a leading risk factor of pancreatic ductal adenocarcinoma (PDAC) that contributes to poor disease prognosis and outcomes. Retrospective studies have identified this link, but interactions surrounding obesity and PDAC are still unclear. Research has shifted to contributions of fibrosis (desmoplasia) on malignancy, which involves increased deposition of collagens and other extracellular matrix (ECM) molecules and increased ECM crosslinking, all of which contribute to increased tissue stiffening. However, fibrotic stiffening is underrepresented as a model feature in current PDAC models. Fibrosis is shared between PDAC and obesity, and can be leveraged for in vitro model design, as current animal obesity models of PDAC are limited in their ability to isolate individual components of fibrosis to study cell behavior. In the current study, methacrylated type I collagen (PhotoCol®) was photo-crosslinked to pathological stiffness levels to recapitulate fibrotic ECM stiffening. PANC-1 cells were encapsulated within PhotoCol®, and the tumor-tissue constructs were prepared to represent normal (healthy) (∼600 Pa) and pathological (∼2000 Pa) tissues. Separately, human mesenchymal stem cells were differentiated into adipocytes representing lean (2D differentiation) and obese fat tissue (3D collagen matrix differentiation), and conditioned media was applied to PANC-1 tumor-tissue constructs. Conditioned media from obese adipocytes showed increased vimentin expression, a hallmark of invasiveness and progression, that was not seen after exposure to media from lean adipocytes or control media. Characterization of the obese adipocyte secretome suggested that some PANC-1 differences may arise from increased interleukin-8 and -10 compared to lean adipocytes. Additionally, high matrix stiffness associated induced an amoeboid morphology in PANC-1 cells that was not present at low stiffness. Amoeboid morphology is an accessory to epithelial-to-mesenchymal transition and is used to navigate complex ECM environments. This plasticity has greater implications for treatment efficacy of metastatic cancers. Overall, this work 1) highlights the importance of investigating PDAC-obesity interactions to study the effects on disease progression and persistence, 2) establishes PhotoCol® as a matrix material that can be leveraged to study amoeboid morphology and invasion in PDAC, and 3) emphasizes the importance of integrating both biophysical and biochemical interactions associated within both pathologies for in vitro PDAC models.

Mechanisms Underlying the Rarity of Skeletal Muscle Cancers.

Skeletal muscle (SKM), despite comprising ~40% of body mass, rarely manifests cancer. This review explores the mechanisms that help to explain this rarity, including unique SKM architecture and function, which prohibits the development of new cancer as well as negates potential metastasis to SKM. SKM also presents a unique immune environment that may magnify the anti-tumorigenic effect. Moreover, the SKM microenvironment manifests characteristics such as decreased extracellular matrix stiffness and altered lactic acid, pH, and oxygen levels that may interfere with tumor development. SKM also secretes anti-tumorigenic myokines and other molecules. Collectively, these mechanisms help account for the rarity of SKM cancer.

Human-derived Tumor-On-Chip model to study the heterogeneity of breast cancer tissue

One of the leading causes that complicate the treatment of some malignancies, including breast cancer, is tumor heterogeneity. In addition to inter-heterogeneity and intra-heterogeneity of tumors that reflect the differences between cancer cell characteristics, heterogeneity in the tumor microenvironment plays a critical role in tumor progression and could be considered an overlooked and a proper target for the effective selection of therapeutic approaches. Due to the difficulty of completely capturing tumor heterogeneity in conventional detection methods, Tumor-on-Chip (TOC) devices with culturing patient-derived spheroids could be an appropriate alternative. In this research, human-derived spheroids from breast cancer individuals were cultured for 6 days in microfluidic devices. To compare TOC data with conventional detection methods, immunohistochemistry (IHC) and ITRAQ data were employed, and various protein expressions were validated using the transcriptomic databases. The behavior of the spheroids in the collagen matrix and the cell viability were monitored over 6 days of culture. IHC and immunocytochemistry (ICC) results revealed that inter and intra-heterogeneity of tumor spheroids are associated with HER2/ER expression. HER2 expression levels revealed a more important biomarker associated with invasion in the 3D culturing of spheroids. The expression levels of CD163 (as a marker for Ma2 macrophages) and CD44 (a marker for cancer stem cells (CSCs)) were also evaluated. Interestingly, the levels of M2a macrophages and CSCs were higher in triple-negative specimens and samples that showed higher migration and invasion. Cell density and extracellular matrix (ECM) stiffness were also important factors affecting the migration and invasion of the spheroids through the matrix. Among these, rigid ECM revealed a more crucial role than cell density. To sum up, these research findings demonstrated that human-derived spheroids from breast cancer specimens in microfluidic devices provide a dynamic condition for predicting tumor heterogeneity in patients, which can help move the field forward for better and more accurate therapeutic strategies.

Intracellular pH dynamics respond to microenvironment stiffening and mediate vasculogenic mimicry through β-catenin.

Dysregulated intracellular pH (pHi) dynamics and an altered tumor microenvironment have emerged as drivers of cancer cell phenotypes. However, the molecular integration between the physical properties of the microenvironment and dynamic intracellular signaling responses remains unclear. Here, we use two metastatic cell models, one breast and one lung, to assess pHi response to varying extracellular matrix (ECM) stiffness. To experimentally model ECM stiffening, we use two tunable-stiffness hydrogel systems: Matrigel and hyaluronic acid (HA) gels, which mimic the increased protein secretion and crosslinking associated with ECM stiffening. We find that single-cell pHi decreases with increased ECM stiffness in both hydrogel systems and both metastatic cell types. We also observed that stiff ECM promotes vasculogenic mimicry (VM), a phenotype associated with metastasis and resistance. Importantly, we show that decreased pHi is both a necessary and sufficient mediator of VM, as raising pHi on stiff ECM reduces VM phenotypes and lowering pHi on soft ECM drives VM. We characterize β-catenin as a pH-dependent molecular mediator of pH-dependent VM, where stiffness-driven changes in β-catenin abundance can be overridden by increased pHi. We uncover a dynamic relationship between matrix stiffness and pHi, thus suggesting pHi dynamics can override mechanosensitive cell responses to the extracellular microenvironment.

A review of the effects of extracellular matrix stiffness on cellular activity

The extracellular matrix is a vital component of a cellular system. It is responsible for several metabolic processes within cells and its properties can have major effects on cellular activity. Recent research has begun to understand the role of physical interactions between the matrix and the cell on cellular growth and repair. In this work, we review the role of the stiffness of the extracellular matrix in determining cellular activity. We first review the effect on stem cell differentiation through the influence of cytoskeletal feedback loops and induced traction stresses. We then discuss the effects of metabolic reprogramming in tumor progression via molecular upregulation of YAP/TAZ and genetic expression. Finally, we highlight the effects of the stiffness gradient dictated by durotaxis and induced hypoxia on the rate of tumor progression. We hope this review sheds light on the major impact that the extracellular matrix has on various biological activities.

Matrix stiffening promotes chondrocyte senescence and the osteoarthritis development through downregulating HDAC3

Extracellular matrix (ECM) stiffening is a typical characteristic of cartilage aging, which is a quintessential feature of knee osteoarthritis (KOA). However, little is known about how ECM stiffening affects chondrocytes and other molecules downstream. This study mimicked the physiological and pathological stiffness of human cartilage using polydimethylsiloxane (PDMS) substrates. It demonstrated that epigenetic Parkin regulation by histone deacetylase 3 (HDAC3) represents a new mechanosensitive mechanism by which the stiffness matrix affected chondrocyte physiology. We found that ECM stiffening accelerated cultured chondrocyte senescence in vitro, while the stiffness ECM downregulated HDAC3, prompting Parkin acetylation to activate excessive mitophagy and accelerating chondrocyte senescence and osteoarthritis (OA) in mice. Contrarily, intra-articular injection with an HDAC3-expressing adeno-associated virus restored the young phenotype of the aged chondrocytes stimulated by ECM stiffening and alleviated OA in mice. The findings indicated that changes in the mechanical ECM properties initiated pathogenic mechanotransduction signals, promoted the Parkin acetylation and hyperactivated mitophagy, and damaged chondrocyte health. These results may provide new insights into chondrocyte regulation by the mechanical properties of ECM, suggesting that the modification of the physical ECM properties may be a potential OA treatment strategy.

Biomechanical Activation of Keloid Fibroblasts Promotes Lysosomal Remodeling and Exocytosis

Keloids are a severe form of scarring for which the underlying mechanisms are poorly understood, and treatment options are limited or inconsistent. While biomechanical forces are potential drivers of keloid scarring, the direct cellular responses to mechanical cues have yet to be defined. The aim of this study was to examine the distinct responses of normal dermal fibroblasts (NDFs) and keloid-derived fibroblasts (KDFs) to changes in extracellular matrix (ECM) stiffness. When cultured on hydrogels mimicking the elasticity of normal or scarred skin, KDFs displayed greater stiffness-dependent increases in cell spreading, F-actin stress fibre formation, and focal adhesion assembly. Elevated acto-myosin contractility in KDFs disrupted the normal mechanical regulation of ECM deposition and conferred resistance myosin inhibitors. Transcriptional profiling identified mechanically-regulated pathways in NDFs and KDFs, including the actin cytoskeleton, Hippo signalling, and autophagy. Further analysis of the autophagy pathway revealed that autophagic flux was intact in both fibroblast populations and depended on acto-myosin contractility. However, KDFs displayed marked changes in lysosome organisation and an increase in lysosomal exocytosis, which was mediated by acto-myosin contractility. Together, these findings demonstrate that KDFs possess an intrinsic increase in cytoskeletal tension, which heightens the response to ECM mechanics and promotes lysosomal exocytosis.

Extracellular matrix stiffness mediates insulin secretion in pancreatic islets via mechanosensitive Piezo1 channel regulated Ca2+ dynamics

The pancreatic islet is surrounded by ECM that provides both biochemical and mechanical cues to the islet β-cell to regulate cell survival and insulin secretion. Changes in ECM composition and mechanical properties drive β-cell dysfunction in many pancreatic diseases. While several studies have characterized changes in islet insulin secretion with changes in substrate stiffness, little is known about the mechanotransduction signaling driving altered islet function in response to mechanical cues. We hypothesized that increasing matrix stiffness will lead to insulin secretion dysfunction by opening the mechanosensitive ion channel Piezo1 and disrupting intracellular Ca2+ dynamics in mouse and human islets. To test our hypothesis, mouse and human cadaveric islets were encapsulated in a biomimetic reverse thermal gel (RTG) scaffold with tailorable stiffness that allows formation of islet focal adhesions with the scaffold and activation of Piezo1 in 3D. Our results indicate that increased scaffold stiffness causes insulin secretion dysfunction mediated by increases in Ca2+ influx and altered Ca2+ dynamics via opening of the mechanosensitive Piezo1 channel. Additionally, inhibition of Piezo1 rescued glucose-stimulated insulin secretion (GSIS) in islets in stiff scaffolds. Overall, our results emphasize the role mechanical properties of the islet microenvironment plays in regulating function. It also supports further investigation into the modulation of Piezo1 channel activity to restore islet function in diseases like type 2 diabetes (T2D) and pancreatic cancer where fibrosis of the peri-islet ECM leads to increased tissue stiffness and islet dysfunction.

Extracellular Matrix Stiffness-Induced Mechanotransduction of Capillarized Liver Sinusoidal Endothelial Cells.

The mechanobiological response mechanism of the fenestrae of liver sinusoidal endothelial cells (LSECs) to the physical stiffness of the extracellular matrix (ECM) remains unclear. We investigated how the mechanical properties of their substrates affect the LSECs' fenestrae by the nitric oxide (NO)-dependent pathway and how they relate to the progression of hepatic sinus capillarization during liver fibrosis. We detected different stiffnesses of ECM in the progress of liver fibrosis (LF) and developed polyacrylamide hydrogel (PAM) substrates to simulate them. Softer stiffness substrates contributed to LSECs maintaining fenestrae phenotype in vitro. The stiffness of liver fibrosis tissue could be reversed in vivo via treatment with anti-ECM deposition drugs. Similarly, the capillarization of LSECs could be reversed by decreasing the ECM stiffness. Our results also indicate that the NO-dependent pathway plays a key regulatory role in the capillarization of ECM-LSECs. Our study reveals ECM-induced mechanotransduction of capillarized LSECs through a NO-dependent pathway via a previously unrevealed mechanotransduction mechanism. The elucidation of this mechanism may offer precise biomechanics-specific intervention strategies targeting liver fibrosis progression.

Stiffness-tunable biomaterials provide a good extracellular matrix environment for axon growth and regeneration.

Neuronal growth, extension, branching, and formation of neural networks are markedly influenced by the extracellular matrix-a complex network composed of proteins and carbohydrates secreted by cells. In addition to providing physical support for cells, the extracellular matrix also conveys critical mechanical stiffness cues. During the development of the nervous system, extracellular matrix stiffness plays a central role in guiding neuronal growth, particularly in the context of axonal extension, which is crucial for the formation of neural networks. In neural tissue engineering, manipulation of biomaterial stiffness is a promising strategy to provide a permissive environment for the repair and regeneration of injured nervous tissue. Recent research has fine-tuned synthetic biomaterials to fabricate scaffolds that closely replicate the stiffness profiles observed in the nervous system. In this review, we highlight the molecular mechanisms by which extracellular matrix stiffness regulates axonal growth and regeneration. We highlight the progress made in the development of stiffness-tunable biomaterials to emulate in vivo extracellular matrix environments, with an emphasis on their application in neural repair and regeneration, along with a discussion of the current limitations and future prospects. The exploration and optimization of the stiffness-tunable biomaterials has the potential to markedly advance the development of neural tissue engineering.

Osr2 functions as a biomechanical checkpoint to aggravate CD8+ T cell exhaustion in tumor

Alterations in extracellular matrix (ECM) architecture and stiffness represent hallmarks of cancer. Whether the biomechanical property of ECM impacts the functionality of tumor-reactive CD8+ Tcells remains largely unknown. Here, we reveal that the transcription factor (TF) Osr2 integrates biomechanical signaling and facilitates the terminal exhaustion of tumor-reactive CD8+ Tcells. Osr2 expression is selectively induced in the terminally exhausted tumor-specific CD8+ Tcell subset by coupled Tcell receptor (TCR) signaling and biomechanical stress mediated by the Piezo1/calcium/CREB axis. Consistently, depletion of Osr2 alleviates the exhaustion of tumor-specific CD8+ Tcells or CAR-T cells, whereas forced Osr2 expression aggravates their exhaustion in solid tumor models. Mechanistically, Osr2 recruits HDAC3 to rewire the epigenetic program for suppressing cytotoxic gene expression and promoting CD8+ Tcell exhaustion. Thus, our results unravel Osr2 functions as a biomechanical checkpoint to exacerbate CD8+ Tcell exhaustion and could be targeted to potentiate cancer immunotherapy.

Early matrix softening contributes to vascular smooth muscle cell phenotype switching and aortic dissection through down-regulation of microRNA-143/145

Thoracic aortic dissection (TAD) is characterized by extracellular matrix (ECM) dysregulation. Aberrations in the ECM stiffness can lead to changes in cellular functions. However, the mechanism by which ECM softening regulates vascular smooth muscle cell (VSMCs) phenotype switching remains unclear. To understand this mechanism, we cultured VSMCs in a soft extracellular matrix and discovered that the expression of microRNA (miR)-143/145, mediated by activation of the AKT signalling pathway, decreased significantly. Furthermore, overexpression of miR-143/145 reduced BAPN-induced aortic softening, switching the VSMC synthetic phenotype and the incidence of TAD in mice. Additionally, high-throughput sequencing of immunoprecipitated RNA indicated that the TEA domain transcription factor 1 (TEAD1) is a common target gene of miR-143/145, which was subsequently verified using a luciferase reporter assay. TEAD1 is upregulated in soft ECM hydrogels in vitro, whereas the switch to a synthetic phenotype in VSMCs decreases after TEAD1 knockdown. Finally, we verified that miR-143/145 levels are associated with disease severity and prognosis in patients with thoracic aortic dissection. ECM softening, as a result of promoting the VSMCs switch to a synthetic phenotype by downregulating miR-143/145, is an early trigger of TAD and provides a therapeutic target for this fatal disease. miR-143/145 plays a role in the early detection of aortic dissection and its severity and prognosis, which can offer information for future risk stratification of patients with dissection.

Heat-inducible CAR-T overcomes adverse mechanical tumor microenvironment in a 3D bioprinted glioblastoma model

Glioblastoma (GBM) presents a significant therapeutic challenge due to the limited efficacy of existing treatments. Chimeric antigen receptor (CAR) T-cell therapy offers promise, but its potential in solid tumors like GBM is undermined by the physical barrier posed by the extracellular matrix (ECM). To address the inadequacies of traditional 2D cell culture, animal models, and Matrigel-based 3D culture in mimicking the mechanical characteristics of tumor tissues, we employed biomaterials and digital light processing-based 3D bioprinting to fabricate biomimetic tumor models with finely tunable ECM stiffness independent of ECM composition. Our results demonstrated that increased material stiffness markedly impeded CAR-T cell penetration and tumor cell cytotoxicity in GBM models. The 3D bioprinted models enabled us to examine the influence of ECM stiffness on CAR-T cell therapy effectiveness, providing a clinically pertinent evaluation tool for CAR-T cell development in stiff solid tumors. Furthermore, we developed an innovative heat-inducible CAR-T cell therapy, effectively overcoming the challenges posed by the stiff tumor microenvironment.

Abstract PO2-02-13: A Mechanical Conditioning gene expression (MeCo) score detects the patients that benefit from neoadjuvant anti-fibrotic therapy in early HER2-negative breast cancer (HER2negBC)

Abstract Background: Extracellular matrix (ECM) stiffness in the tumor microenvironment is associated with aggressive features that lead to adverse clinical outcomes, including metastasis. Mechanical conditioning (MC) is the phenomenon whereby cancer cells that have grown in a fibrotic-like stiff ECM acquire aggressive features which can be maintained after metastasis in distant sites (Cell Rep.; 35:109293). MeCo Score is a gene expression signature that measures the response to ECM stiffness in early tumors which is associated with subsequent metastasis. We sought to study whether MeCo Score is associated with pathologic complete response (pCR) in early HER2negBC, and whether anti-fibrotic treatment with nintedanib (a multikinase VEGFR1-3, FGFR1-3 and PDGFRA/B inhibitor approved for idiopathic lung fibrosis) is able to mitigate the impact of MC. Methods: Baseline tumor samples from patients (N=130) from a randomized phase II clinical trial (Clin Cancer Res 23: 1432-42) in early HER2negBC trial comparing weekly paclitaxel monotherapy (85 mg/m2 x 12 courses; N=65; Arm A) versus the same treatment plus nintedanib (150 mg p.o. bid, continuous schedule; Arm B) were analyzed by RNAseq. MeCo Score was computed as previous described (Cell Rep.; 35:109293), and intrinsic PAM50 subtypes were determined using the R package genefu. Clinical, demographic and pathologic variables were gathered for all patients; pCR was graded according to the Residual Cancer Burden (RCB) score, RCB=0. Regression models were built using the variables RCB, T, N, G, age, intrinsic subtype, Ki67, MeCo Score and treatment arm. Bi-variate correlations were analyzed with the Pearson coefficient. All statistic tests were two-sided. Results: MeCo Score was successfully determined in 76 women (50% from each arm); the average score was 0.34 (range: 0.052 – 0.617). Of them, 48.6% were Luminal A, 34.2% Luminal B, 14.5% Basal-like, and 2.7% Normal according to PAM50. Median age was 49 y.o. (range: 30.6 – 79.2); tumor size distribution was T1:3.0%; T2:68.2%; T3:24.3%; T4: 4.5%; 47% of the patients were N0, 47% were N1 and 6% were N2. Most patients (85%) were ER and/or PR positive; 15% were “triple-negative”. Histological grade distribution was G1/G2/G3: 15%/62%/23%. The average Ki67 was 28.7% (range: 1%-98%). pCR was 10.8% and 4.5% in arms A and B. The average MeCo Score by intrinsic subtype was 0.26 (LumA); 0.43 (LumB) and 0.38 (Basal), LumB vs LumA P< 0.001; LumB vs Basal: P=0.037. We did not find correlation between MeCo Score and G, T, N, age or Ki67, neither in the Luminal subgroup or the whole cohort. In Arm A, patients with high MeCo Score (i.e., above median) had a 0% pCR rate compared to 10.5% pCR rate for those with low MeCo Score (P=0.035); however, in the nintedanib arm (Arm B) no differences were found: 10.5% versus 11.1% pCR rate in high versus low MeCo Score (P=0.4). Conclusions: Among early HER2negBC, Luminal B cases display the highest MeCo Scores. MeCo Score was independent of all classic prognostic variables. High MeCo Score is associated with lack of pCR to neoadjuvant paclitaxel monotherapy, but concurrent administration of the anti-fibrotic agent nintedanib rescues this association. Citation Format: Miguel Quintela-Fandino, Adam Watson, Adam Grant, Gus Mouneimne, Maria J. Bueno, Silvana Mouron. A Mechanical Conditioning gene expression (MeCo) score detects the patients that benefit from neoadjuvant anti-fibrotic therapy in early HER2-negative breast cancer (HER2negBC) [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-02-13.