3D Tumor Models Research Articles

Application and progress of 3D tumor models in breast cancer.

Due to its high heterogeneity and significant impact on women's health globally, breast cancer necessitates robust preclinical models to understand tumor biology and guide personalized treatment strategies. Three-dimensional (3D) in vitro tumor models hold immense promise in this regard. These tumor models not only mimic the spatial structure and growth environment of tumors in vivo, but also retain the pathological and genetic characteristics of solid tumors. This fidelity makes them powerful tools for accelerating advancements in fundamental research and translational medicine. The diversity, modularity, and efficacy of 3D tumor models are driving a biotechnological revolution. As these technologies become increasingly sophisticated, 3D tumor models are poised to become powerful weapons in the fight against breast cancer. This article expounds on the progress made in utilizing 3D tumor models for breast cancer research.

Inhibition of OXPHOS induces metabolic rewiring and reduces hypoxia in murine tumor models

IntroductionTumor hypoxia is a feature of many solid malignancies and is known to cause radio resistance. In recent years it has become clear that hypoxic tumor regions also foster an immunosuppressive phenotype and are involved in immunotherapy resistance. It has been proposed that reducing the tumors’ oxygen consumption will result in an increased oxygen concentration in the tissue and improve radio- and immunotherapy efficacy. The aim of this study is to investigate the metabolic rewiring of cancer cells by pharmacological attenuation of oxidative phosphorylation (OXPHOS) and subsequently reduce tumor hypoxia. Material and methodsThe metabolic effects of three OXPHOS inhibitors IACS-010759, atovaquone and metformin were explored by measuring oxygen consumption rate, extra cellular acidification rate, and [18F]FDG uptake in 2D and 3D cell culture. Tumor cell growth in 2D cell culture and hypoxia in 3D cell culture were analyzed by live cell imaging. Tumor hypoxia and [18F]FDG uptake in vivo following treatment with IACS-010759 was determined by immunohistochemistry and ex vivo biodistribution respectively. ResultsIn vitro experiments show that tumor cell metabolism is heterogeneous between different models. Upon OXPHOS inhibition, metabolism shifts from oxygen consumption through OXPHOS towards glycolysis, indicated by increased acidification and glucose uptake. Inhibition of OXPHOS by IACS-010759 treatment reduced diffusion limited tumor hypoxia in both 3D cell culture and in vivo. Although immune cell presence was lower in hypoxic areas compared with normoxic areas, it is not altered following short term OXPHOS inhibition. DiscussionThese results show that inhibition of OXPHOS causes a metabolic shift from OXPHOS towards increased glycolysis in 2D and 3D cell culture. Moreover, inhibition of OXPHOS reduces diffusion limited hypoxia in 3D cell culture and murine tumor models. Reduced hypoxia by OXPHOS inhibition might enhance therapy efficacy in future studies. However, caution is warranted as systemic metabolic rewiring can cause adverse effects.

Linking Metastatic Potential and Viscoelastic Properties of Breast Cancer Spheroids via Dynamic Compression and Relaxation in Microfluidics.

The growth and invasion of solid tumors are associated with changes in their viscoelastic properties, influenced by both internal cellular factors and physical forces in the tumor microenvironment. Due to the lack of a comprehensive investigation of tumor tissue viscoelasticity, the relationship between such physical properties and cancer malignancy remains poorly understood. Here, the viscoelastic properties of breast cancer spheroids, 3D (in vitro) tumor models, are studied in relation to their metastatic potentials by imposing controlled, dynamic compression within a microfluidic constriction, and subsequently monitoring the relaxation of the imposed deformation. By adopting a modified Maxwell model to extract viscoelastic properties from the compression data, the benign (MCF-10A) spheroids are found to have higher bulk elastic modulus and viscosity compared to malignant spheroids (MCF-7 and MDA-MB-231). The relaxation is characterized by two timescales, captured by a double exponential fitting function, which reveals a similar fast rebound for MCF-7 and MCF-10A. Both the malignant spheroids exhibit similar long-term relaxation and display residual deformation. However, they differ significantly in morphology, particularly in intercellular movements. These differences between malignant spheroids are demonstrated to be linked to their cytoskeletal organization, by microscopic imaging of F-actin within the spheroids, together with cell-cell adhesion strength.

Paper Spray Ionization Mass Spectrometry Coupled with Paper-Based Three-Dimensional Tumor Model for Rapid Metabolic Gradient Profiling.

The tumor microenvironment (TME), especially with its complicated metabolic characteristics, will dynamically affect the proliferation, migration, and drug response of tumor cells. Rapid metabolic analysis brings out a deeper understanding of the TME, while the susceptibility and environmental dependence of metabolites extremely hinder real-time metabolic profiling since the TME is easily disrupted. Here, we directly integrated paper spray ionization mass spectrometry with a paper-based three-dimensional (3D) tumor model, realizing the rapid capture of metabolic gradients. The entire procedure, from sample preparation to mass spectrometry detection, took less than 4 min, which was able to provide metabolic results close to real time and contributed to understanding the real metabolic processes. At present, our method successfully detected 160 metabolites; notably, over 40 significantly gradient metabolites were revealed across the six layers of the paper-based 3D tumor model. At least 22 gradient metabolites were reported to be associated with cell viability. This strategy was powerful enough to rapidly profile metabolic gradients of a paper-based 3D tumor model for revealing cell viability changes from a metabolomics perspective.

Biobased hydrogel bioinks of pectin, nanocellulose and lysozyme nanofibrils for the bioprinting of A375 melanoma cell-laden 3D in vitro platforms

Melanoma is one of the most aggressive types of skin cancer, and the need for advanced platforms to study this disease and to develop new treatments is rising. 3D bioprinted tumor models are emerging as advanced tools to tackle these needs, with the design of adequate bioinks being a fundamental step to address this challenging process. Thus, this work explores the synergy between two biobased nanofibers, nanofibrillated cellulose (NFC) and lysozyme nanofibrils (LNFs), to create pectin nanocomposite hydrogel bioinks for the 3D bioprinting of A375 melanoma cell-laden living constructs. The incorporation of LNFs (5, 10 or 15 wt%) on a Pectin-NFC suspension originates inks with enhanced rheological properties (shear viscosity and yield point) and proper shear-thinning behavior. The crosslinked hydrogels mimic the stiffness of melanoma, being stable under physiological and cell-culture conditions, and non-cytotoxic towards A375 melanoma cells. P-NFC-LNFs (10 %) reveals good printability (Pr = 0.89) and printing accuracy (51 ± 2 %), and when loaded with A375 cells (3 × 106 cells mL−1) the bioink originates 3D-constructs with high cell viability (92 ± 1 %) after 14 days. The potential of the constructs as in vitro models is corroborated by a drug-screening test with doxorubicin, where cells within the model displayed high sensitivity to the drug.

NIR Enhanced pH-Responsive Microneedles for Synergetic Therapy of Melanoma.

Melanoma has emerged as a significant threat to human life and health. Microneedle (MN)-mediated transdermal drug delivery (TDD) has garnered attention in melanoma treatment for bypassing the first-pass effect. However, the propensity of melanoma to metastasize presents substantial challenges for MN mediated local treatment. Developing systemic therapies, such as immunotherapy in combination with TDD, is crucial for achieving effective melanoma treatment. Herein, a polyvinyl alcohol (PVA) MN-mediated multifunctional TDD system, designated MN@PDA@1-MT/CUR/DOX@HA (MN@PMCDH), was developed for synergetic chemotherapy/photothermal/immunotherapy of melanoma. PMCDH nanomedicines penetrate deep skin layers through MNs, accumulate at tumor sites guided by hyaluronic acid (HA), and selectively release drugs in response to the acidic tumor microenvironment and near-infrared (NIR) stimulation. Released curcumin (CUR) significantly enhances the efficacy of photothermal therapy (PTT) and chemotherapy, as well as improves the induction of immunogenic cell death (ICD) by increasing melanoma sensitivity to polydopamine (PDA)-mediated photothermal effects and doxorubicin (DOX). Moreover, the incorporation of 1-methyltryptophan (1-MT) to reverse the tumor immunosuppressive microenvironment can further enhance the effects of immunotherapy. In vitro studies revealed that the MN@PMCDH system can effectively induce ICD and inhibit tumor cell growth. Additionally, remarkable deep tumor cell inhibition effects are also achieved in 3D tumor models.

Hyaluronan-Based Hydrogels for 3D Modeling of Tumor Tissues.

Although routine two-dimensional (2D) cell culture techniques have advanced basic cancer research owing to their simplicity, cost-effectiveness, and reproducibility, they have limitations that necessitate the development of advanced three-dimensional (3D) tumor models that better recapitulate the tumor microenvironment. Various biomaterials have been used to establish these 3D models, enabling the study of cancer cell behavior within different matrices. Hyaluronic acid (HA), a key component of the extracellular matrix (ECM) in tumor tissues, has been widely studied and employed in the development of multiple cancer models. This review first examines the role of HA in tumors, including its function as an ECM component and regulator of signaling pathways that affect tumor progression. It then explores HA-based models for various cancers, focusing on HA as a central component of the 3D matrix and its mobilization within the matrix for targeted studies of cell behavior and drug testing. The tumor models discussed included those for breast cancer, glioblastoma, fibrosarcoma, gastric cancer, hepatocellular carcinoma, and melanoma. The review concludes with a discussion of future prospects for developing more robust and high-throughput HA-based models to more accurately mimic the tumor microenvironment and improve drug testing. Impact Statement This review underscores the transformative potential of hyaluronic acid (HA)-based hydrogels in developing advanced tumor models. By exploring HA's dual role as a critical extracellular matrix component and a regulator of cancer cell dynamics, we highlight its unique contributions to replicating the tumor microenvironment. The recent advancements in HA-based models provide new opportunities for more accurate studies of cancer cell behavior and drug responses. Looking ahead, these innovations pave the way for high-throughput, biomimetic platforms that could revolutionize drug testing and accelerate the discovery of effective cancer therapies.

Three-dimensional breast cancer tumor models based on natural hydrogels: a review.

Breast cancer is the most common cancer in women and one of the deadliest cancers worldwide. According to the distribution of tumor tissue, breast cancer can be divided into invasive and non-invasive forms. The cancer cells in invasive breast cancer pass through the breast and through the immune system or systemic circulation to different parts of the body, forming metastatic breast cancer. Drug resistance and distant metastasis are the main causes of death from breast cancer. Research on breast cancer has attracted extensive attention from researchers. In vitro construction of tumor models by tissue engineering methods is a common tool for studying cancer mechanisms and anticancer drug screening. The tumor microenvironment consists of cancer cells and various types of stromal cells, including fibroblasts, endothelial cells, mesenchymal cells, and immune cells embedded in the extracellular matrix. The extracellular matrix contains fibrin proteins (such as types I, II, III, IV, VI, and X collagen and elastin) and glycoproteins (such as proteoglycan, laminin, and fibronectin), which are involved in cell signaling and binding of growth factors. The current traditional two-dimensional (2D) tumor models are limited by the growth environment and often cannot accurately reproduce the heterogeneity and complexity of tumor tissues in vivo. Therefore, in recent years, research on three-dimensional (3D) tumor models has gradually increased, especially 3D bioprinting models with high precision and repeatability. Compared with a 2D model, the 3D environment can better simulate the complex extracellular matrix components and structures in the tumor microenvironment. Three-dimensional models are often used as a bridge between 2D cellular level experiments and animal experiments. Acellular matrix, gelatin, sodium alginate, and other natural materials are widely used in the construction of tumor models because of their excellent biocompatibility and non-immune rejection. Here, we review various natural scaffold materials and construction methods involved in 3D tissue-engineered tumor models, as a reference for research in the field of breast cancer.

Abstract A027: Enhancing pancreatic cancer chemotherapy through photochemical internalisation

Abstract Introduction Pancreatic cancer is the deadliest common cancer. Its poor prognosis is associated with its complex biology and lack of early detection strategies. Although much progress has been made in finding more effective therapeutic strategies, pancreatic tumours remain difficult to treat. A major reason why chemotherapy is relatively ineffective against this type of cancer is inefficient delivery of drugs to their target sites combined with multidrug resistance. Our aim is to use the technique called Photochemical Internalisation (PCI), a light-triggered intracellular drug delivery method which combines low dose Photodynamic Therapy (PDT) with chemotherapy, to induce efficient cytosolic delivery of therapeutic compounds to their specific subcellular targets. Methodology Treatment outcomes using saporin (a type I ribosome-inactivating protein) and gemcitabine as monotherapies, or in combination with a light-activated photosensitizer (the porphyrin TPPS2a), were thoroughly analysed in both established pancreatic cancer cell lines and patient-derived xenograft (PDX) models. The efficacy of these treatments was assessed in both 2D and 3D tumour models through various cell viability assays, including MTT, crystal violet staining, and neutral red staining. Additionally, molecular techniques such as western blotting, protein arrays, and flow cytometry were employed to determine the effects on cell viability and to elucidate changes in signalling pathways leading to the therapeutic response. Results In this study, we observed minimal cytotoxicity induced by saporin, gemcitabine or TPPS2a + light monotherapies. However, PCI synergistically enhanced saporin and gemcitabine cytotoxicity (p<0.001) using very low concentrations in all tumour models. Moreover, we determined the mechanism of cell death triggered by these protocols. PCI induced endosomal disruption following light excitation and apoptosis leading to caspases 3/7 activation, in a time dependent manner. Conclusions Overall, our findings demonstrate the potential of PCI to enhance the efficacy of cancer chemotherapy for pancreatic cancer using lower doses than in monotherapy and open a new window for future translational studies. By using low doses of light therapy, we have found a promising avenue for future studies that could eventually lead to better treatments for this challenging disease. Citation Format: Maria Rosado, Ismahan Mahamed, Andres Garcia-Sampedro, Sandy MacRoberts, Pål Selbo, Stephen Pereira, Pilar Acedo. Enhancing pancreatic cancer chemotherapy through photochemical internalisation [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A027.

Stat3 Inhibitors TTI-101 and SH5-07 Suppress Bladder Cancer Cell Survival in 3D Tumor Models.

Bladder cancer (BCa) is one of the most lethal genitourinary malignancies owing to its propensity for recurrence and poor survival. The biochemical pathway, signal transducer and activator of transcription 3 (STAT3), has gained significance as a molecular pathway that promotes proliferation, invasion, and chemoresistance. In this study, we explored the targeting of STAT3 with TTI-101 and SH5-07 in BCa and elucidated the mechanisms in three-dimensional (3D) spheroid and organoid models. We optimized the growth of spheroids from human, rat, and mouse BCa cell lines (J82, NBT-II, and MB49 respectively) and organoids from BBN (N-butyl-N-(4-hydroxybutyl)-nitrosamine)-induced rat bladder tumors. Cell viability was assessed using MTT and trypan blue assays. Intracellular ATP production, ROS production, and calcium AM (CA)/EtBr staining were used to measure the spheroid and organoid inhibition and mitochondrial function. Western blot analysis was performed to evaluate the pharmacodynamic markers involved in cell proliferation, apoptosis, cancer stem cells (CSCs), and STAT3 signaling in BCa. We found that targeting STAT3 (using TTI-101 and SH5-07) significantly reduced the proliferation of BCa spheroids and organoids, which was accompanied by decreased expression of pSTAT3, Cyclin D1, and PCNA. Our data also demonstrated that treatment with STAT3 inhibitors induced ROS production and cell death in BCa spheroids and organoids. STAT3 inhibition-induced cell death was associated with the activation of caspase 3/7 and PARP cleavage. Moreover, TTI-101 and SH5-07 target cancer stem cells by downregulating the expression of CD44 and CD133 in 3D models. This study provides the first evidence for the prevention of BCa with small-molecule inhibitors TTI-101 and SH5-07 via suppression of CSCs and STAT3 signaling.

Ultrasensitive Dual Fluorophore-Conjugated Carbon Dots for Intracellular pH Sensing in 3D Tumor Models.

The dysregulation of pH has been linked to the onset of chronic conditions, such as cancer and neurological diseases. Consequently, the development of a highly sensitive tool for intracellular pH sensing is imperative to investigate the interplay between pH and the biochemical changes accompanying disease pathogenesis. Here, we present the development of a ratiometric fluorescent nanoprobe, NpRhoDot, designed for precisely measuring pH levels. We demonstrate its efficacy in sensitively reporting intracellular pH in monolayer A549 lung cancer cells, primary fibroblast cells, and 3D tumor spheroids derived from the DLD-1 colorectal adenocarcinoma cell line. NpRhoDot leverages a novel design, where stable carbon dots are functionalized with a pH-responsive ratiometric fluorescent probe comprising a naphthalimide-rhodamine moiety, NpRho1. This design confers NpRhoDot with the high pH sensitivity characteristics of organic fluorescent probes, along with excellent photostability up to 1 h and biocompatibility of carbon dots. Through one-photon and two-photon fluorescence microscopy, we validate the reliability of NpRhoDot for biosensing intracellular pH in monolayer and three-dimensional tumor models from pH 4 to 7.

Assessing Drug Uptake and Response Differences in 2D and 3D Cellular Environments Using Stimulated Raman Scattering Microscopy.

The architecture of cell culture, two-dimensional (2D) versus three-dimensional (3D), significantly impacts various cellular factors, including cell-cell interactions, nutrient and oxygen gradients, metabolic activity, and gene expression profiles. This can result in different cellular responses during cancer drug treatment, with 3D-cultured cells often exhibiting higher resistance to chemotherapeutic drugs. While various genetic and proteomic analyses have been employed to investigate the underlying mechanisms of this increased resistance, complementary techniques that provide experimental evidence of spatial molecular profiling data are limited. Stimulated Raman scattering (SRS) microscopy has demonstrated its capability to measure both intracellular drug uptake and growth inhibition. In this work, we applied three-band (C-D, C-H, and fingerprint regions) SRS imaging to 2D and 3D cell cultures and performed a comparative analysis of drug uptake and response with the goal of understanding whether the difference in drug uptake explains the drug resistance in 3D culture compared to 2D. Our investigations revealed that despite similar intracellular drug levels in 2D and 3D A549 cells during lapatinib treatment, the growth of 3D spheroids was less impacted, supporting an enhanced drug tolerance in the 3D microenvironment. We further elucidated drug penetration patterns and the resulting heterogeneous cellular responses across different spheroid layers. Additionally, we investigated the role of the extracellular matrix in modulating drug delivery and cell response and discovered that limited drug penetration in 3D could also contribute to lower drug response. Our study provides valuable insights into the intricate mechanisms of increased drug resistance in 3D tumor models during cancer drug treatments.

Next-generation 3D tumor modeling: A microfluidic platform with biocompatible red carbon dots for live cell imaging in co-cultured elongated spheroid tumor model

Co-culture spheroids mimic tumor architecture more accurately than traditional 2D cell cultures, but non-invasive, long-term tracking of live cells within these 3D models remains a challenge. This study addresses this critical need by developing a novel approach for live cell imaging in U-87/HUF co-culture spheroids. We introduce water-soluble, biocompatible red carbon dots (R-CDs) exhibiting exceptional stability and brightness (21% quantum yield) specifically designed for imaging within these 3D models. Furthermore, we designed a microfluidic chip with ellipsoid-shaped microwells to efficiently generate two distinct co-culture spheroid types: direct mixing and core-shell. R-CDs enabled non-invasive tracking of U-87 cancer cell location within these 3D models demonstrating their efficacy for long-term monitoring of live cells in cancer research. This R-CD and microfluidic technology has the potential to accelerate cancer drug discovery by enabling live cell studies in 3D tumor models.

Development of A 3D Lung Tumor Model For Cell Death Assessment in Microwave Ablation

AbstractPrimary lung cancer is the leading cause of cancer‐related death worldwide. Recent statistics estimate 1.8 million new cases and 1.6 million deaths per year. Despite significant advances in surgery, chemotherapy, and radiotherapy, the 5‐year survival rate remains at just 18%. Thermal ablation is an established treatment that is safe and effective. It refers to the local application of controlled heat to induce irreversible cell injury and ultimately tumor apoptosis and necrosis. This approach is particularly suitable for the treatment of small tumors in patients who are not suitable for surgery. However, the lack of suitable benchtop models to assess ablation devices complicates prognostic prediction and the development of effective treatment strategies. In this context, the development of a 3D lung tumor model is presented using a cancer cell‐laden hydrogel composed of 2% hyaluronic acid–tyramine (HA‐TA) and 0.5% agarose, resulting in a tumor mimetic of clinically relevant size. This innovative model provides a valuable platform for testing ablation devices and provides insights that can significantly improve prognostic ability and refine treatment approaches for lung cancer patients.

Estimation of the injection criteria for magnetic hyperthermia therapy based on tumor morphology

Intratumoral multi-injection strategy enhances the efficacy of magnetic nanoparticle hyperthermia therapy (MNPH). In this study, criteria for the selection of injections and their location depending on the tumor shape/geometry are developed. The developed strategy is based on the thermal dosimetry results of different invasive 3D tumor models during MNPH simulation. MNPH simulations are conducted on physical tumor tissue models encased within healthy tissue. The tumor shapes are geometrically divided into a central tumor region containing maximum tumor volume and a peripheral tumor portion protruding in any random direction. The concepts of core and invasive radius are used to geometrically divide the tumor volume. Primary & secondary injections are used to inject MNP fluid into these respective tumor regions based on the invasiveness of the tumor. The optimization strategy is devised based on the zone of influence of primary & secondary injection. Results indicate that the zone of influence of secondary injection lies between 0.7 and 0.8 times the radial distance between the center of the tumor core and branch node point (extreme far endpoint on the invasive tumor surface). Additionally, the multi-injection strategy is more effective when the protrusion volume exceeds 10% of the total volume. The proposed algorithm is used to devise multi-injection strategies for arbitrarily shaped tumors and will assist in pre-planning magnetic nanoparticle hyperthermia therapy.

Tumor morphology evaluation using 3D-morphometric features of renal masses.

To assess the correlation between the general (gender, age, and maximum tumor size) and 3D morphotopometric features of the renal tumor node, following the MSCT data post-processing, and the tumor histological structure; to propose an equation allowing for kidney malignancy assessment based on general and morphometric features. In total, 304 patients with unilateral solitary renal neoplasms underwent laparoscopic (retroperitoneoscopic) or robotic partial or radical nephrectomy. Before the procedure, kidney contrast-enhanced MSCT followed by the tumor 3D-modeling was performed. 3D model of the kidney tumor, and its morphotopometric features, and histological structure were analyzed. The morphotopometric ones include the side of the lesion, location by segments, the surface where the tumor, the depth of the tumor invasion into the kidney, and the shape of tumor. Out of 304 patients, 254 (83.6%) had malignant kidney tumors and 50 (16.4%) benign kidney tumors. In total, 231 patients, out of 254 (90.9%) were assessed for the degree of malignant tumor differentiation. Malignant tumors were more frequent in men than in women (p < 0.001). Mushroom-shaped tumors were the most common shapes among benign renal masses (35.2%). The most common malignant kidney tumors had spherical with a partially uneven surface (27.6%), multinodular (tuberous (27.2%)), and spherical with a conical base (24.8%) shapes. Logistic regression model enabled the development of prognostic equation for tumor malignancy prediction ("low" or "high"). The univariate analysis revealed the correlation only between high differentiation (G1) and a spherical tumor with a conical base (p = 0.029). The resulting logistic model, based on the analysis of such predictors as gender and form of kidney lesions, demonstrated a large share (87.6%) of correct predictions of the kidney tumor malignancy.

3D bioprinting of an in vitro hepatoma microenvironment model: Establishment, evaluation, and anticancer drug testing

Tumor behavior, including its response to treatments, is influenced by interactions between mesenchymal and malignant cells, as well as their spatial arrangement. To study tumor biology and evaluate anticancer drugs, accurate 3D tumor models are essential. Here, we developed an in vitro biomimetic hepatoma microenvironment model by combining an extracellular matrix (3DM-7721). Initially, the internal grid structure, composed of 10/6 % GelMA/gelatin loaded with SMMC-7721 cells, was printed using 3D bioprinting. The external component consisted of fibroblasts and human umbilical vein endothelial cells loaded with 10/3 % GelMA/gelatin. A control model (3DP-7721) lacked external cell loading. GelMA/gelatin hydrogels provided robust structural support and biocompatibility. The SMMC-7721 cells in the 3DM-7721 model exhibit superior tumor-associated gene expression and proliferation characteristics when compared to the 3DP-7721 model. Furthermore, the 3DM-7721 type exhibited increased resistance to anticancer agents. SMMC-7721 cells in the 3DM-7721 model exhibit significant tumorigenicity in nude mice. The 3DM-7721 model group showed pathological characteristics of malignant tumors, with a high degree of deterioration, and a significant positive correlation between malignant tumor-related gene pathways. This high-fidelity 3DM-7721 tumor microenvironment model is invaluable for studying tumor progression, devising effective treatment strategies, and discovering drugs. Statement of significance1.A hepatoma microenvironment model (3DM-7721) incorporating an extracellular matrix that is designed to establish solid tumor models in vitro.2.This model contains a 3D-printed internal tumor component of liver cancer cells loaded with 10/6 % gelatin methacrylate (GelMA)/gelatin (the 3DP-7721 model) and an external extracellular-matrix component of fibroblasts and human umbilical vein endothelial cells loaded with 10/3 % GelMA/gelatin.3.The 3DM-7721 model outperforms the 3DP-7721 model in terms of the SMMC-7721 cell proliferation characteristics and tumorigenicity, tumor-related gene expression, anticancer drug resistance, and malignant tumor characteristics.

Surface-Enhanced Raman Scattering Monitoring of Tryptophan Dynamics in 3D Pancreatic Tumor Models.

In the intricate landscape of the tumor microenvironment, both cancer and stromal cells undergo rapid metabolic adaptations to support their growth. Given the relevant role of the metabolic secretome in fueling tumor progression, its unique metabolic characteristics have gained prominence as potential biomarkers and therapeutic targets. As a result, rapid and accurate tools have been developed to track metabolic changes in the tumor microenvironment with high sensitivity and resolution. Surface-enhanced Raman scattering (SERS) is a highly sensitive analytical technique and has been proven efficient toward the detection of metabolites in biological media. However, profiling secreted metabolites in complex cellular environments such as those in tumor-stroma 3D in vitro models remains challenging. To address this limitation, we employed a SERS-based strategy to investigate the metabolic secretome of pancreatic tumor models within 3D cultures. We aimed to monitor the immunosuppressive potential of stratified pancreatic cancer-stroma spheroids as compared to 3D cultures of either pancreatic cancer cells or cancer-associated fibroblasts, focusing on the metabolic conversion of tryptophan into kynurenine by the IDO-1 enzyme. We additionally sought to elucidate the dynamics of tryptophan consumption in correlation with the size, temporal evolution, and composition of the spheroids, as well as assessing the effects of different drugs targeting the IDO-1 machinery. As a result, we confirm that SERS can be a valuable tool toward the optimization of cancer spheroids, in connection with their tryptophan metabolizing capacity, potentially allowing high-throughput spheroid analysis.

Selective anticancer effects of plasma-activated saline in 3D tumor model co-culturing of normal cells and cancer cells

Compared with conventional two-dimensional (2D) cell culture model, the 3D tumor model constructed in vitro is better representative of the tumor microenvironment in vivo. Here, we proposed the utilization of 3D tumor model of co-cultured cancer cells and normal cells to evaluate the selective anticancer effects of cold atmospheric plasma-activated saline (PAS), and expected to provide more precise information about PAS-tumor interactions. By cell sorting, we clarified that A375 melanoma cells and HaCaT normal skin cells purified from the 3D multicellular tumor model differ in sensitivity and responsiveness to PAS compared to the 2D culture model. And during the optimization of PAS treatment parameters, we further found that A375 cells were almost completely killed while HaCaT cells were still present in large numbers after 5 d of certain PAS treatment. Our experiment innovatively carries out the selective study of plasma technology in 3D co-culture system and provides a theoretical basis for further clinical and practical applications of PAS.

A 3D Pancreatic Cancer Model with Integrated Optical Sensors for Noninvasive Metabolism Monitoring and Drug Screening.

A distinct feature of pancreatic ductal adenocarcinoma (PDAC) is a prominent tumor microenvironment (TME) with remarkable cellular and spatial heterogeneity that meaningfully impacts disease biology and treatment resistance. The dynamic crosstalk between cancer cells and the dense stromal compartment leads to spatially and temporally heterogeneous metabolic alterations, such as acidic pH that contributes to drug resistance in PDAC. Thus, monitoring the extracellular pH metabolic fluctuations within the TME is crucial to predict and to quantify anticancer drug efficacy. Here,a simple and reliable alginate-based 3D PDAC model embedding ratiometric optical pH sensors and cocultures of tumor (AsPC-1) and stromal cells for simultaneously monitoring metabolic pH variations and quantify drug response is presented. By means of time-lapse confocal laser scanning microscopy (CLSM) coupled with a fully automated computational analysis, the extracellular pH metabolic variations are monitored and quantified over time during drug testing with gemcitabine, folfirinox, and paclitaxel, commonly used in PDAC therapy. In particular, the extracellular acidification is more pronounced after drugs treatment, resulting in increased antitumor effect correlated with apoptotic cell death. These findings highlight the importance of studying the influence of cellular metabolic mechanisms on tumor response to therapy in 3D tumor models, this being crucial for the development of personalized medicine approaches.