Engineered nascent living human tissues with unit programmability.

Leveraging human cells as materials precursors is a promising approach for fabricating living materials with tissue-like functionalities and cellular programmability. Here we describe a set of cellular units with metabolically engineered glycoproteins that allow cells to tether together to function as macrotissue building blocks and bioeffectors. The generated human living materials, termed as Cellgels, can be rapidly assembled in a wide variety of programmable three-dimensional configurations with physiologically relevant cell densities (up to 108 cells per cm3), tunable mechanical properties and handleability. Cellgels inherit the ability of living cells to sense and respond to their environment, showing autonomous tissue-integrative behaviour, mechanical maturation, biological self-healing, biospecific adhesion and capacity to promote wound healing. These living features also enable the modular bottom-up assembly of multiscale constructs, which are reminiscent of human tissue interfaces with heterogeneous composition. This technology can potentially be extended to any human cell type, unlocking the possibility for fabricating living materials that harness the intrinsic biofunctionalities of biological systems.

Strategies to decouple cell micro-scale and macro-scale environments for designing multifunctional biomimetic tissues.

The regulation of cellular behavior within a three-dimensional (3D) environment to execute a specific function remains a challenge in the field of tissue engineering. In native tissues, cells and matrices are arranged into 3D modular units, comprising biochemical and biophysical signals that orchestrate specific cellular activities. Modular tissue engineering aims to emulate this natural complexity through the utilization of functional building blocks with unique stimulation features. By adopting a modular approach and using well-designed biomaterials, cellular microenvironments can be effectively decoupled from their macro-scale surroundings, enabling the development of engineered tissues with enhanced multifunctionality and heterogeneity. We overview recent advancements in decoupling the cellular micro-scale niches from their macroenvironment and evaluate the implications of this strategy on cellular and tissue functionality.

Exploring the potential of all-aqueous immiscible systems for preparing complex biomaterials and cellular constructs.

All-aqueous immiscible systems derived from liquid-liquid phase separation of incompatible hydrophilic agents such as polymers and salts have found increasing interest in the biomedical and tissue engineering fields in the last few years. The unique characteristics of aqueous interfaces, namely their low interfacial tension and elevated permeability, as well as the non-toxic environment and high water content of the immiscible phases, confer to these systems optimal qualities for the development of biomaterials such as hydrogels and soft membranes, as well as for the preparation of in vitro tissues derived from cellular assembly. Here, we overview the main properties of these systems and present a critical review of recent strategies that have been used for the development of biomaterials with increased levels of complexity using all-aqueous immiscible phases and interfaces, and their potential as cell-confining environments for micropatterning approaches and the bioengineering of cell-rich structures. Importantly, due to the relatively recent emergence of these areas, several key design considerations are presented, in order to guide researchers in the field. Finally, the main present challenges, future directions, and adaptability to develop advanced materials with increased biomimicry and new potential applications are briefly evaluated.

Photocrosslinkable microgels derived from human platelet lysates: injectable biomaterials for cardiac cell culture.

Cardiovascular diseases are a major global cause of morbidity and mortality, and they are often characterized by cardiomyocytes dead that ultimately leads to myocardial ischemia (MI). This condition replaces functional cardiac tissue with fibrotic scar tissue compromising heart function. Injectable systems for the in situ delivery of cells or molecules to assist during tissue repair have emerged as promising approaches for tissue engineering, particularly for myocardial repair. Methacryloyl platelet lysates (PLMA) have been employed for constructing full human-based 3D cell culture matrices and demonstrated potential for xeno-free applications. In this study, we propose using PLMA to produce microparticles (MPs) serving as anchors for cardiac and endothelial cells and ultimately as injectable systems for cardiac tissue repair. The herein reported PLMA MPs were produced by droplet microfluidics and showed great properties for cell attachment. More importantly, it is possible to show the capacity of PLMA MPs to serve as cell microcarriers even in the absence of animal-derived serum supplementation in the culture media.

Regulation of cell fate by cell imprinting approach in vitro.

Cell culture-based technologies are widely utilized in various domains such as drug evaluation, toxicity assessment, vaccine and biopharmaceutical development, reproductive technology, and regenerative medicine. It has been demonstrated that pre-adsorption of extracellular matrix (ECM) proteins including collagen, laminin and fibronectin provide more degrees of support for cell adhesion. The purpose of cell imprinting is to imitate the natural topography of cell membranes by gels or polymers to create a reliable environment for the regulation of cell function. The results of recent studies show that cell imprinting is a tool to guide the behavior of cultured cells by controlling their adhesive interactions with surfaces. Therefore, in this review we aim to compare different cell cultures with the imprinting method and discuss different cell imprinting applications in regenerative medicine, personalized medicine, disease modeling, and cell therapy.

Modeling 3D Tumor Invasiveness to Modulate Macrophage Phenotype in a Human-Based Hydrogel Platform.

The immune system is a pivotal player in determining tumor fate, contributing to the immunosuppressive microenvironment that supports tumor progression. Considering the emergence of biomaterials as promising platforms to mimic the tumor microenvironment, human platelet lysate (PLMA)-based hydrogel beads are proposed as 3D platforms to recapitulate the tumor milieu and recreate the synergistic tumor-macrophage communication. Having characterized the biomaterial-mediated pro-regenerative macrophage phenotype, an osteosarcoma spheroid encapsulated into a PLMA hydrogel bead is explored to study macrophage immunomodulation through paracrine signaling. The culture of PLMA-Tumor beads on the top of a 2D monolayer of macrophages reveals that tumor cells triggered morphologic and metabolic adaptations in macrophages. The cytokine profile, coupled with the upregulation of gene and protein anti-inflammatory biomarkers clearly indicates macrophage polarization toward an M2-like phenotype. Moreover, the increased gene expression of chemokines identified as pro-tumoral environmental regulators suggest a tumor-associated macrophage phenotype, exclusively stimulated by tumor cells. This pro-tumoral microenvironment is also found to enhance tumor invasiveness ability and proliferation. Besides providing a robust in vitro immunomodulatory tumor model that faithfully recreates the tumor-macrophage interplay, this human-based platform has the potential to provide fundamental insights into immunosuppressive signaling and predict immune-targeted response.

Embedded 3D Bioprinting for Engineering Miniaturized In Vitro Tumor Models.

Embedded extrusion 3D bioprinting is a rapidly emerging additive manufacturing methodology that provides a precise spatial deposition of synthetic or natural-origin low-viscosity bioinks during the extrusion printing process. Such a strategy has to date unlocked the freeform extrusion biofabrication of complex micro-to-macro-scale living architectures for numerous applications, including tissue engineering and in vitro disease modeling. In this chapter, we describe a suspension bioprinting methodology leveraging a continuous viscoelastic biopolymer supporting bath functionalized with divalent calcium cations to enable a rapid processing of user-defined bioinks toward architecturally complex 3D in vitro tumor models. This highly simple and cost-effective viscoelastic supporting bath enables a full freeform biofabrication of cell-laden 3D tumor-mimetic architectures that exhibit structural stability in culture post-printing. The cytocompatibility of the supporting bath, its ease of removal from biofabricated living constructs, and its adaptability for processing different ECM-mimetic bioinks open avenues for multi-scale fabrication of numerous types of physiomimetic 3D tumor models for preclinical screening of candidate therapeutics.

Magneto-Enzymatic Microgels for Precise Hydrogel Sculpturing.

The inclusion of hollow channels in tissue-engineered hydrogels is crucial for mimicking the natural physiological conditions and facilitating the delivery of nutrients and oxygen to cells. Although bio-fabrication techniques provide diverse strategies to create these channels, many require sophisticated equipment and time-consuming protocols. Herein, collagenase, a degrading agent for methacrylated gelatin hydrogels, and magnetic nanoparticles (MNPs) are combined and processed into enzymatically active spherical structures using a straightforward oil bath emulsion methodology. The generated microgels are then used to microfabricate channels within biomimetic hydrogels via a novel sculpturing approach that relied on the precise coupling of protein-enzyme pairs (for controlled local degradation) and magnetic actuation (for directional control). Results show that the sculpting velocity can be tailored by adjusting the magnetic field intensity or concentration of MNPs within the microgels. Additionally, varying the magnetic field position or microgel size generated diverse trajectories and channels of different widths. This innovative technology improves the viability of encapsulated cells through enhanced medium transport, outperforming non-sculpted hydrogels and offering new perspectives for hydrogel vascularization and drug/biomolecule administration. Ultimately, this novel concept can help design fully controlled channels in hydrogels or soft materials, even those with complex tortuosity, in a single wireless top-down biocompatible step.

Advances in Cell-Rich Inks for Biofabricating Living Architectures.

Advancing biofabrication toward manufacturing living constructs with well-defined architectures and increasingly biologically relevant cell densities is highly desired to mimic the biofunctionality of native human tissues. The formulation of tissue-like, cell-dense inks for biofabrication remains, however, challenging at various levels of the bioprinting process. Promising advances have been made toward this goal, achieving relatively high cell densities that surpass those found in conventional platforms, pushing the current boundaries closer to achieving tissue-like cell densities. On this focus, herein the overarching challenges in the bioprocessing of cell-rich living inks into clinically grade engineered tissues are discussed, as well as the most recent advances in cell-rich living ink formulations and their processing technologies are highlighted. Additionally, an overview of the foreseen developments in the field is provided and critically discussed.

Encapsulation of pristine and silica-coated human adipose-derived mesenchymal stem cells in gelatin colloidal hydrogels for tissue engineering and bioprinting applications.

Colloidal gels assembled from gelatin nanoparticles (GNPs) as particulate building blocks show strong promise to solve challenges in cell delivery and biofabrication, such as low cell survival and limited spatial retention. These gels offer evident advantages to facilitate cell encapsulation, but research on this topic is still limited, which hampers our understanding of the relationship between the physicochemical and biological properties of cell-laden colloidal gels. Human adipose-derived mesenchymal stem cells were successfully encapsulated in gelatin colloidal gels and evaluated their mechanical and biological performance over 7 days. The cells dispersed well within the gels without compromising gel cohesiveness, remained viable, and spread throughout the gels. Cells partially coated with silica were introduced into these gels, which increased their storage moduli and decreased their self-healing capacity after 7 days. This finding demonstrates the ability to modulate gel stiffness by incorporating cells partially coated with silica, without altering the solid content or introducing additional particles. Our work presents an efficient method for cell encapsulation while preserving gel integrity, expanding the applicability of colloidal hydrogels for tissue engineering and bioprinting. Overall, our study contributes to the design of improved cell delivery systems and biofabrication techniques.

Contribution of non-steroidal anti-inflammatory drugs to breast cancer treatment: In vitro and in vivo studies.

Chronic inflammation plays a crucial role in carcinogenesis. High levels of serum prostaglandin E2 and tissue overexpression of cyclooxygenase-2 (COX-2) have been described in breast, urinary, colorectal, prostate, and lung cancers as being involved in tumor initiation, promotion, progression, angiogenesis, and immunosuppression. Non-steroidal anti-inflammatory drugs (NSAIDs) are prescribed for several medical conditions to not only decrease pain and fever but also reduce inflammation by inhibiting COX and its product synthesis. To date, significant efforts have been made to better understand and clarify the interplay between cancer development, inflammation, and NSAIDs with a view toward addressing their potential for cancer management. This review provides readers with an overview of the potential use of NSAIDs and selective COX-2 inhibitors for breast cancer treatment, highlighting pre-clinical in vitro and in vivo studies employed to evaluate the efficacy of NSAIDs and their use in combination with other antineoplastic drugs.

Acidochromic Free-Standing Multilayered Chitosan-Pyranoflavylium/Alginate Membranes toward Food Smart Packaging Applications.

Food smart packaging has emerged as a promising technology to address consumer concerns regarding food conservation and food safety. In this context, we report the rational design of azide-containing pyranoflavylium-based pH-sensitive dye for subsequent click chemistry conjugation toward a chitosan-modified alkyne. The chitosan-pyranoflavylium conjugate was characterized by infrared (ATR-FTIR), ultraviolet-visible (UV-vis), nuclear magnetic resonance (NMR) spectroscopies, and dynamic light scattering (DLS), as well as its thermodynamic parameters related to their pH-dependent chromatic features. The fabrication of thin-films through electrostatic-driven layer-by-layer (LbL) assembly technology was first screened by quartz crystal microbalance with dissipation monitoring (QCM-D) onto gold substrates, and then free-standing (FS) multilayered membranes from polypropylene substrate were obtained using a homemade automatic dipping robot. The membranes' characterization included morphology analysis and thickness evaluation, assessed by scanning electron microscopy (SEM), pH-responsive color change performance tests using buffer solutions at different pH levels, and biogenic amines-enriched model solutions, demonstrating the feasibility and effectiveness of the chitosan-pyranoflavylium/alginate biomembranes for food spoilage monitoring. This work provides insights toward the development of innovative pH-responsive smart biomaterials for advanced and sustainable technological packaging solutions, which could significantly contribute to ensuring food safety and quality, while reducing food waste.

In-Bath 3D Printing of Anisotropic Shape-Memory Cryogels Functionalized with Bone-Bioactive Nanoparticles.

Cryogels exhibit unique shape memory with full recovery and structural stability features after multiple injections. These constructs also possess enhanced cell permeability and nutrient diffusion when compared to typical bulk hydrogels. Volumetric processing of cryogels functionalized with nanosized units has potential to widen their biomedical applications, however this has remained challenging and relatively underexplored. In this study, we report a novel methodology that combines suspension 3D printing with directional freezing for the fabrication of nanocomposite cryogels with configurable anisotropy. When compared to conventional bulk or freeze-dried hydrogels, nanocomposite cryogel formulations exhibit excellent shape recovery (>95%) and higher pore connectivity. Suspension printing, assisted with a prechilled metal grid, was optimized to induce anisotropy. The addition of calcium- and phosphate-doped mesoporous silica nanoparticles into the cryogel matrix enhanced bioactivity toward orthopedic applications without hindering the printing process. Notably, the nanocomposite 3D printed cryogels exhibit injectable shape memory while also featuring a lamellar topography. The fabrication of these constructs was highly reproducible and exhibited potential for a cell-delivery injectable cryogel with no cytotoxicity to human-derived adipose stem cells. Hence, in this work, it was possible to combine a gravity defying 3D printed methodology with injectable and controlled anisotropic macroporous structures containing bioactive nanoparticles. This methodology ameliorates highly tunable injectable 3D printed anisotropic nanocomposite cryogels with a user-programmable degree of structural complexity.

A scaffold vaccine to promote tumor antigen cross-presentation via sustained toll-like receptor-2 (TLR2) activation.

Cancer vaccination holds great promise for cancer treatment, but its effectiveness is hindered by suboptimal activation of CD8+ cytotoxic T lymphocytes, which are potent effectors to mediate anti-tumor immune responses. A possible solution is to switch antigen-presenting cells to present tumor antigens via the major histocompatibility complex class I (MHC-I) to CD8+ T cells - a process known as cross-presentation. To achieve this goal, we develop a three-dimensional (3D) scaffold vaccine to promote antigen cross-presentation by persisted toll-like receptor-2 (TLR2) activation after one injection. This vaccine comprises polysaccharide frameworks that "hook" TLR2 agonist (acGM) via tunable hydrophobic interactions and forms a 3D macroporous scaffold via click chemistry upon subcutaneous injection. Its retention-and-release of acGM enables sustained TLR2 activation in abundantly recruited dendritic cells in situ, inducing intracellular production of reactive oxygen species (ROS) in optimal kinetics that crucially promotes efficient antigen cross-presentation. The scaffold loaded with model antigen ovalbumin (OVA) or tumor specific antigen can generate potent immune responses against lung metastasis in B16-OVA-innoculated wild-type mice or spontaneous colorectal cancer in transgenic ApcMin/+ mice, respectively. Notably, it requires neither additional adjuvants nor external stimulation to function and can be adjusted to accommodate different antigens. The developed scaffold vaccine may represent a new, competent tool for next-generation personalized cancer vaccination.

Human Platelet Lysate-Derived Nanofibrils as Building Blocks to Produce Free-Standing Membranes for Cell Self-Aggregation.

Amyloid-like fibrils are garnering keen interest in biotechnology as supramolecular nanofunctional units to be used as biomimetic platforms to control cell behavior. Recent insights into fibril functionality have highlighted their importance in tissue structure, mechanical properties, and improved cell adhesion, emphasizing the need for scalable and high-kinetics fibril synthesis. In this study, we present the instantaneous and bulk formation of amyloid-like nanofibrils from human platelet lysate (PL) using the ionic liquid cholinium tosylate as a fibrillating agent. The instant fibrillation of PL proteins upon supramolecular protein-ionic liquid interactions was confirmed from the protein conformational transition toward cross-ß-sheet-rich structures. These nanofibrils were utilized as building blocks for the formation of thin and flexible free-standing membranes via solvent casting to support cell self-aggregation. These PL-derived fibril membranes reveal a nanotopographically rough surface and high stability over 14 days under cell culture conditions. The culture of mesenchymal stem cells or tumor cells on the top of the membrane demonstrated that cells are able to adhere and self-organize in a three-dimensional (3D) spheroid-like microtissue while tightly folding the fibril membrane. Results suggest that nanofibril membrane incorporation in cell aggregates can improve cell viability and metabolic activity, recreating native tissues' organization. Altogether, these PL-derived nanofibril membranes are suitable bioactive platforms to generate 3D cell-guided microtissues, which can be explored as bottom-up strategies to faithfully emulate native tissues in a fully human microenvironment.

Encapsulated Mesenchymal Stromal Cells as Cyclic Providers of Immunomodulatory Secretomes: A Living on-Demand Delivery System.

The stimulation of mesenchymal stromal cells (MSCs) with inflammatory molecules is often used to boost their therapeutic effect. Prolonged exposure to inflammatory molecules has been explored to improve their action because MSCs therapies seem to be improved transiently with such stimuli. However, the possibility of cyclically stimulating MSCs to recover their optimized therapeutic potential is still to be elucidated, although the efficacy of cell-based therapies may be dependent on the ability to readapt to the relapse pathological conditions. Here, the response of MSCs, encapsulated in alginate hydrogels and cultured for 22 d, is explored using three different regimes: single, continuous, and intermittent stimulation with IFNγ. Exposure to IFNγ leads to a decrease in the secretion of IL-10, which is cyclically countered by IFNγ weaning. Conditioned media collected at different stages of pulsatile stimulation show an immunomodulatory potential toward macrophages, which directly correlates with IL-10 concentration in media. To understand whether the correlation between cyclic stimulation of MSCs and other biological actions can be observed, the effect on endothelial cells is studied, showcasing an overall modest influence on tube formation. Overall, the results describe the response of encapsulated MSCs to unusual pulsatile simulation regimens, exploring encapsulated MSCs as a living on-demand release system of tailored secretomes with recoverable immunomodulatory action.

Photoacoustic processing of decellularized extracellular matrix for biofabricating living constructs.

The diverse biomolecular landscape of tissue-specific decellularized extracellular matrix (dECM) biomaterials provides a multiplicity of bioinstructive cues to target cells, rendering them highly valuable for various biomedical applications. However, the isolation of dECM biomaterials entails cumbersome xenogeneic enzymatic digestions and also additional inactivation procedures. Such, increases processing time, increments costs and introduces residues of non-naturally present proteins in dECM formulations that remain present even after inactivation. To overcome these limitations, herein we report an innovative conjugation of light and ultrasound-mediated dECM biomaterial processing for fabricating dECM biomaterials. Such approach gathers on ultrasound waves to facilitate dECM-in-liquid processing and visible light photocrosslinking of tyrosine residues naturally present in dECM biomaterials. This dual step methodology unlocked the in-air production of cell laden dECM hydrogels or programmable dECM hydrogel spherical-like beads by using superhydrophobic surfaces. These in-air produced units do not require any additional solvents and successfully supported both fibroblasts and breast cancer cells viability upon encapsulation or surface seeding. In addition, the optimized photoacoustic methodology also enabled a rapid formulation of dECM biomaterial inks with suitable features for biofabricating volumetrically defined living constructs through embedded 3D bioprinting. The biofabricated dECM hydrogel constructs supported cell adhesion, spreading and viability for 7 days. Overall, the implemented photoacoustic processing methodology of dECM biomaterials offers a rapid and universal strategy for upgrading their processing from virtually any tissue. STATEMENT OF SIGNIFICANCE: Leveraging decellularized extracellular matrix (dECM) as cell instructive biomaterials has potential to open new avenues for tissue engineering and in vitro disease modelling. The processing of dECM remains however, lengthy, costly and introduces non-naturally present proteins in the final biomaterials formulations. In this regard, here we report an innovative light and ultrasound two-step methodology that enables rapid dECM-in-liquid processing and downstream photocrosslinking of dECM hydrogel beads and 3D bioprinted constructs. Such photoacoustic based processing constitutes a universally applicable method for processing any type of tissue-derived dECM biomaterials.

Comparative analysis of aligned and random amniotic membrane-derived cryogels for neural tissue repair.

The ordered arrangement of cells and extracellular matrix facilitates the seamless transmission of electrical signals along axons in the spinal cord and peripheral nerves. Therefore, restoring tissue geometry is crucial for neural regeneration. This study presents a novel method using proteins derived from the human amniotic membrane, which is modified with photoresponsive groups, to produce cryogels with aligned porosity. Freeze-casting was used to produce cryogels with longitudinally aligned pores, while cryogels with randomly distributed porosity were used as the control. The cryogels exhibited remarkable injectability and shape-recovery properties, essential for minimally invasive applications. Different tendencies in proliferation and differentiation were evident between aligned and random cryogels, underscoring the significance of the scaffold's microstructure in directing the behaviour of neural stem cells (NSC). Remarkably, aligned cryogels facilitated extensive cellular infiltration and migration, contrasting with NSC cultured on isotropic cryogels, which predominantly remained on the scaffold's surface throughout the proliferation experiment. Significantly, the proliferation assay demonstrated that on day 7, the aligned cryogels contained eight times more cells compared to the random cryogels. Consistent with the proliferation experiments, NSC exhibited the ability to differentiate into neurons within the aligned scaffolds and extend neurites longitudinally. In addition, differentiation assays showed a four-fold increase in the expression of neural markers in the cross-sections of the aligned cryogels. Conversely, the random cryogels exhibited minimal presence of cell bodies and extensions. The presence of synaptic vesicles on the anisotropic cryogels indicates the formation of functional synaptic connections, emphasizing the importance of the scaffold's microstructure in guiding neuronal reconnection.

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.

Advances in screening hyperthermic nanomedicines in 3D tumor models.

Hyperthermic nanomedicines are particularly relevant for tackling human cancer, providing a valuable alternative to conventional therapeutics. The early-stage preclinical performance evaluation of such anti-cancer treatments is conventionally performed in flat 2D cell cultures that do not mimic the volumetric heat transfer occurring in human tumors. Recently, improvements in bioengineered 3D in vitro models have unlocked the opportunity to recapitulate major tumor microenvironment hallmarks and generate highly informative readouts that can contribute to accelerating the discovery and validation of efficient hyperthermic treatments. Leveraging on this, herein we aim to showcase the potential of engineered physiomimetic 3D tumor models for evaluating the preclinical efficacy of hyperthermic nanomedicines, featuring the main advantages and design considerations under diverse testing scenarios. The most recent applications of 3D tumor models for screening photo- and/or magnetic nanomedicines will be discussed, either as standalone systems or in combinatorial approaches with other anti-cancer therapeutics. We envision that breakthroughs toward developing multi-functional 3D platforms for hyperthermia onset and follow-up will contribute to a more expedited discovery of top-performing hyperthermic therapies in a preclinical setting before their in vivo screening.