Bridging biomaterials and immunotherapy: hydrogel strategies for next-generation CAR-T cell treatment

Abstract

Breast cancer remains a leading cause of cancer-associated mortality worldwide, with an increasing incidence among younger women, particularly in developing regions and immigrant populations. Although advances in diagnostics and targeted therapies have improved patient outcomes, the search for more effective and less toxic treatments continues. Chimeric antigen receptor (CAR) T-cell therapy has transformed cancer immunotherapy by harnessing the body’s own immune system to fight malignancy, yet its full potential in solid tumors is still being realized. Meanwhile, hydrogel-based biomaterials have emerged as versatile platforms capable of local immune modulation, sustained cytokine delivery, and enhanced T-cell activation within the tumor microenvironment. By bridging materials science with tumor immunology, this evolving field offers a new perspective on how to direct immune activity precisely where it is needed. Combining hydrogel technology with CAR-T cell therapy represents a promising strategy to strengthen antitumor responses and improve treatment precision and safety.

Introduction

With over 2.3 million new cases and about 700,000 deaths annually, breast cancer stands as both the most diagnosed malignancy and one of the leading causes of cancer-related mortality in women worldwide, underscoring its persistent global health burden [1, 2]. Although significant progress has been made in screening, surgery, chemotherapy, radiotherapy, and targeted agents; conventional treatments often fail in advanced or metastatic disease [3, 4]. Particularly in triple-negative breast cancer (TNBC), treatment options are limited due to the absence of actionable molecular targets and the rapid emergence of therapeutic resistance [5, 6]. Moreover, the immunologically “cold” nature of breast cancer, characterized by low T cell infiltration and high immunosuppression, further limits the success of immune checkpoint inhibitors and other standard immunotherapies [7].

Adoptive cell therapies, particularly those involving chimeric antigen receptor (CAR)- T cells, have transformed the landscape of hematologic malignancies by inducing durable remissions in patients with otherwise refractory disease. However, their clinical efficacy in solid tumors such as breast cancer remains restricted [8–10]. Major challenges that promote T cell exhaustion and dysfunction include poor T cell trafficking and retention within the tumor, antigen heterogeneity, and a highly suppressive tumor microenvironment (TME) [11, 12]. Additionally, systemic delivery of CAR-T cells can provoke significant toxicities which highlights the need for spatial precision to concentrate immune responses at the tumor site. These obstacles have driven efforts to design new strategies for localized and sustained cell activity in solid tumors [13].

To address these challenges, biomaterial-based approaches, particularly hydrogel systems, have gained increasing attention [14]. Hydrogels are highly hydrated, cross-linked polymeric matrices that can be designed to carry cells, cytokines, or immunomodulatory compounds and release them in a controlled, site-specific manner [15]. Their properties, such as porosity, mechanical strength, and biodegradability, can be adjusted to meet the needs of different therapies [16]. In the context of cancer immunotherapy, injectable hydrogels offer a unique opportunity to create a supportive microenvironment for CAR-T cells, enhance their survival, expansion, and effector function while shield them from hostile elements within the TME. Moreover, hydrogel depots can be formulated to respond to local stimuli, such as pH or enzymes, which makes it possible to release immune agents only when needed and helps avoid off-target effects in healthy tissues [17–19].

This review first examines the tumor microenvironment and the processes that drive its progression, then explores CAR-T cell therapy and the challenges it faces in treating breast cancer. Finally, we highlight how combining CAR-T cells with hydrogel-based strategies provides an innovative way to boost therapeutic effectiveness while minimizing side effects, paving the way for the next generation of more precise and patient-centered immunotherapies in breast cancer.

Aggressive breast cancer in young women requires immunotherapy solutions

While breast cancer has traditionally been associated with postmenopausal women, recent epidemiological evidence reveals a disturbing shift toward earlier onset. Increasingly, breast cancer is being diagnosed in women under 45 years of age—many of whom are still within their reproductive years and have not completed family planning [20–22]. This younger subgroup often develops biologically aggressive forms of the disease, most notably triple-negative and HER2-positive breast cancers, which are known for their high proliferation rates, early metastatic potential, and resistance to standard therapies [23, 24]. Compared to their older counterparts, young breast cancer patients often harbor tumors enriched with immunosuppressive components of the tumor microenvironment (TME), including elevated levels of regulatory T cells (Tregs), tumor-associated macrophages (TAMs), and myeloid-derived suppressor cells (MDSCs). These cellular players contribute to immune escape and reduce responsiveness to both chemotherapy and immunotherapy [25, 26]. Although immunotherapy has shown encouraging results in cancers like melanoma and lung cancer, its effectiveness in breast cancer is still inconsistent. Researchers are increasingly interested in testing these approaches in younger patients, whose immune systems might behave differently and could be more responsive to treatment. In particular, strategies such as immune checkpoint inhibitors and adoptive cell therapies are currently under investigation in clinical trials for aggressive breast cancer subtypes that tend to occur more frequently in this age group [25].

To improve outcomes for this rising high-risk population, treatment strategies must integrate immunologic profiling with age-specific tumor biology. Future efforts should focus on optimizing combination regimens and enhancing localized immune activation, particularly through innovative delivery systems such as biomaterial-assisted platforms. Such precision approaches may overcome current limitations and offer a new therapeutic horizon for young women facing aggressive breast cancer.

The immunoediting paradigm in breast cancer: an immunologic overview.

Within this context, the concept of cancer immunoediting provides valuable insights into the complex relationship between the immune system and tumors. It highlights how immunity can both restrain and shape tumor development. In breast cancer, immunoediting proceeds through three overlapping stages—elimination, equilibrium, and escape—illustrating the dynamic interplay between malignant cells and the host immune system. This ongoing dialogue influences tumor immunogenicity, drives disease progression, and ultimately impacts clinical outcomes [23, 27].

Mechanisms of immune surveillance: elimination

In the elimination phase, the innate and adaptive immune systems act in concert to detect and eradicate nascent malignant cells. Innate immune cells such as natural killer (NK) cells, macrophages, and dendritic cells (DCs) are instrumental in the early detection of transformed breast epithelial cells. NK cells mediate cytotoxicity against tumor cells via the release of perforin and granzymes, while also produce interferon-gamma (IFN-γ), promote antigen presentation and shape adaptive immune responses. Dendritic cells capture tumor antigens and migrate to tumor-draining lymph nodes to prime tumor-specific CD8⁺ cytotoxic T lymphocytes (CTLs), which are critical for cancer cells death. This is further augmented by CD4⁺ helper T cells, which enhance CTL expansion and macrophage activation through cytokine secretion (e.g., IL-2, IFN-γ). Macrophages, through nitric oxide and Tumor Necrosis Factor Alpha (TNF-α) production, contribute to direct tumor cell killing and immune amplification. The successful orchestration of these responses can result in complete tumor eradication before clinical manifestation [28–31].

Tumor equilibrium

When tumor cells evade complete elimination, they may persist in a state of immune-mediated equilibrium, during which immune pressure constrains tumor outgrowth without fully eradicating it. This phase, which may span months to years, is governed primarily by adaptive immune components; most notably CD8⁺ T cells and IFN-γ signaling. The sustained immune surveillance exerts selective pressure on tumor cells, facilitates the emergence of less immunogenic variants through antigen loss, MHC class I downregulation, or resistance to cytotoxic mechanisms. In breast cancer, equilibrium is particularly relevant in the context of minimal residual disease or dormancy, where micro metastases remain clinically undetectable yet immunologically restrained [32–37].

Immune escape

Ultimately, the escape phase is characterized by the emergence of tumor cell populations that have acquired or selected for mechanisms to subvert immune recognition and destruction. Breast cancer cells may upregulate immune checkpoint molecules such as Programmed Death-Ligand 1 (PD-L1), recruit immunosuppressive cell populations including regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs), and secrete inhibitory cytokines like Transforming Growth Factor Beta (TGF-β) and Interleukin 10 (IL-10). All of these processes result in the establishment of the tumor microenvironment (TME). Concurrently, defects in antigen processing and presentation pathways, metabolic competition (e.g., via suppression of T-cell activation and promotion of Treg by cytokine secretion), and hypoxia dampens T-cell functionality. These adaptations facilitate uncontrolled tumor growth, immune evasion, and metastatic dissemination and hallmarks of clinical breast cancer progression [38–40].

Chimeric antigen receptor T cell (CAR-T cell)

Breast cancer cells, especially aggressive subtypes like TNBC, often evade immune surveillance and limit chemotherapy and radiotherapy efficacy [5–7]. CAR-T therapy, successful in hematologic cancers, is being explored for solid tumors, including breast cancer, but faces challenges such as tumor heterogeneity, immunosuppressive microenvironments, poor T cell trafficking, off-tumor toxicity, and cytokine release syndrome [9, 13, 41, 42]. CAR-T cells are engineered T cells that recognize tumor-associated antigens (TAAs) like HER2, MUC1, Trop2, and EGFR independently of MHC, allowing them to bypass immune evasion and selectively eliminate tumor cells [43–48]. Advances from first-generation to fourth- and fifth-generation CARs, incorporating co-stimulatory domains (CD28, 4-1BB) and inducible cytokines (IL-7, IL-15, IL-21), have improved T cell persistence, proliferation, cytokine secretion, and resistance to exhaustion, enhancing efficacy within the immunosuppressive tumor microenvironment [49–52].

CAR-T cells limitations and challenges

The immunosuppressive tumor microenvironment (TME)

The tumor microenvironment (TME) strongly affects CAR-T therapy in solid tumors like breast cancer [53]. Unlike blood cancers, solid tumors have an immunosuppressive environment with stromal cells, TAMs, MDSCs, Tregs, and dense ECM, which hinder T-cell migration, survival, and cytotoxicity [54–57]. Hypoxia and factors like TGF-β, PD-1, and CTLA-4 further suppress immunity, though PD-1/PD-L1 blockade can restore T-cell activity [58–60]. CAFs also promote T-cell exclusion via SDF-1α (CXCR4 ligand) [61–64]. Metabolic reprogramming of CAR-T cells, enhancing pyruvate- and MPC-dependent oxidative phosphorylation (OXPHOS), can rejuvenate exhausted CD8 + T cells and boost cytotoxicity and antitumor immunity [59]. Strategies to overcome TME barriers include dominant-negative TGF-β receptors, chemokine receptors like CXCR2 [65–67], and combining CAR-T cells with checkpoint inhibitors or ECM-degrading enzymes such as heparinase [68, 69]. Mesothelin-targeted CAR-T cells show promise in TNBC models with these approaches (Fig. 1).

Fig. 1.

The immunosuppressive tumor microenvironment (TME) comprises tumor cells, tumor-associated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs), and cancer-associated fibroblasts (CAFs). These components secrete inhibitory cytokines and stromal cell–derived factor-1α (SDF-1α), promoting CAR-T cell dysfunction and exhaustion. Exhausted CAR-T cells exhibit increased expression of inhibitory receptors, including PD-1 and CXCR4, which interact with PD-L1 and SDF-1α within the TME, thereby reinforcing immunosuppressive signaling. Created in BioRender. shojaei, S. (2026) https://BioRender.com/kz4l2an

Barriers to CAR-T cell trafficking and tumor infiltration

A major limitation of CAR-T therapy in breast cancer is poor trafficking and infiltration into the tumor parenchyma [70, 71]. The immunosuppressive TME presents physical and molecular barriers, including a dense ECM rich in collagen and hyaluronan, abnormal vasculature, and high interstitial fluid pressure, all hindering T-cell extravasation and migration [72, 73]. Additionally, chemokine profiles in tumors often do not match CAR-T cell receptors [74].

The CXCR3 pathway, expressed on activated CD4 + Th1, CD8 + T, NK, and NKT cells, guides immune cells to tumors via ligands CXCL9–11, enhancing infiltration and correlating with better outcomes [68, 75]. CAR-T cells can also be engineered to express chemokine receptors matching tumor signals; for example, CXCR2 enables migration toward CXCL1, improving infiltration, tumor regression, and survival in preclinical models[76–78]. Conversely, blocking inhibitory axes like CXCL12–CXCR4, which restricts T-cell entry and promotes tumor growth, can enhance CAR-T efficacy, with several CXCR4 antagonists in clinical trials [79, 80]. Similarly, GD2-CAR T cells engineered with CCR2b showed ~ tenfold higher tumor accumulation and improved antitumor activity by following CCL2 gradients (Table 1) [81, 82].

Table 1.

Major challenges of CAR-T cell and hydrogel-based combination strategies

CAR-T Cell Challenges	Hydrogel-based Combination Solutions
I. Immunosuppressive tumor microenvironment (TME)	I. Reshaping the Tumor Microenvironment with Hydrogel-Enhanced CAR-T Cell Therapy
II. Barriers to CAR-T Cell Trafficking and Tumor Infiltration	II. Improving CAR-T Cell Trafficking and Cytotoxicity in Breast Cancer via Hydrogel Scaffolds
III. Antigen Heterogeneity and Escape Mechanisms in Solid Tumors	III. Smart Hydrogel Systems for Precision-Controlled CAR-T Cell Expansion and Targeted Breast Cancer Therapy
IV. On-Target, Off-Tumor Toxicities	IV. Enhancing Tumor Specificity to Prevent Off-Tumor Toxicity
V. Cytokine released toxicity	V. Strategic Modulation of CAR-T Cell Activation to Mitigate Cytokine Release Syndrome (CRS)

Antigen heterogeneity and escape mechanisms in solid tumors

Antigen heterogeneity is a major barrier to effective CAR-T therapy [83]. In breast cancer, inter- and intratumoral heterogeneity create diverse antigenic profiles within and between tumors [84, 85]. Intratumoral heterogeneity means a single tumor may contain cell subpopulations with distinct genetic and molecular features, so a monoclonal CAR-T targeting a single TAA (e.g., HER2 or MUC1) may miss cells that lack or underexpress the antigen [86]. The TME can further reinforce heterogeneity by selecting for antigen-low or -negative clones, promoting immune evasion [82, 87].

This dynamic antigen landscape limits CAR-T durability compared with hematologic malignancies where target expression (e.g., CD19) is more uniform [88, 89]. Heterogeneity also affects immunosuppressive ligand expression (e.g., PD-L1) and cytokine profiles, modulating CAR-T function and persistence [90, 91]. Antigen escape—downregulation or mutation of the target following CAR-T pressure—can occur via transcriptional repression, alternative splicing, post-translational modifications, or genomic deletion, leading to loss of CAR engagement [92, 93]. In breast cancer, non-uniform TAA expression makes this especially problematic [94–96].

To overcome these challenges, strategies include dual-targeting or logic-gated CARs and combination therapies with checkpoint inhibitors, broadening immune coverage and preventing escape [96–98]. Single-cell mapping of heterogeneity is essential for rational next-generation CAR-T design (Table 1).

On-target, off-tumor toxicities

On-target/off-tumor toxicity is a major challenge in CAR-T therapy, especially in solid tumors like breast cancer, where many antigens are also expressed at low levels on normal tissues [99, 100]. This occurs when CAR-T cells correctly recognize their target (on-target) on non-malignant cells (off-tumor), causing tissue damage [101, 102]. For instance, HER2-targeted CAR-T can affect pulmonary epithelium and cardiac tissue, with fatal pulmonary toxicity reported due to low HER2 expression on lung cells [103–105]. CAR-T activation triggers perforin/granzyme and Fas–FasL–mediated killing, along with inflammatory cytokine release (IFN-γ, TNF-α, IL-6, IL-10), intensifying local and systemic toxicity [106, 107].

Activation thresholds depend on CAR affinity, antigen density, epitope accessibility, and co-stimulatory domains [108, 109]. High-affinity CARs are potent but more likely to target low-level antigens on healthy tissue. Unlike endogenous TCRs, CARs lack thymic negative selection, increasing off-tumor risk [110]. Strategies to improve specificity include dual-antigen or AND-gate CARs, tunable CARs with inducible signaling, and synNotch receptor systems that prime CAR expression only in response to tumor-specific antigens, thereby limiting activity in normal tissues [111–115] (Table 1).

When the immune system overreacts: cytokine release syndrome

Cytokine Release Syndrome (CRS) is a key immunologic complication of CAR-T therapy, arising from robust immune activation after CAR-T engagement with tumor antigens [110, 116]. While less frequent in breast cancer than hematologic malignancies, CAR-Ts targeting solid tumor antigens (e.g., mesothelin, HER2, MUC1, c-Met) raise concern for CRS [10, 117]. CAR-T activation via CD3ζ and co-stimulatory domains (CD28, 4-1BB) triggers rapid secretion of cytokines (IL-6, IL-1, IL-10, IFN-γ, TNF-α), activating monocytes/macrophages and creating a feed-forward loop that drives systemic inflammation, endothelial activation, and vascular leakage (Fig. 2) [118, 119].

Fig. 2.

Cytokine release syndrome (CRS) results from excessive immune activation following CAR-T cell recognition of tumor antigens. Antigen-dependent CAR signaling drives rapid release of pro-inflammatory cytokines, including IL-6, IL-1, IFN-γ, and TNF-α. These mediators activate monocytes and macrophages, amplifying systemic inflammation and leading to endothelial dysfunction, vascular leakage and organ damage. Created in BioRender. shojaei, S. (2026) https://BioRender.com/db070lh

In TNBC and other aggressive breast tumors, high tumor burden amplifies CRS due to extensive immune-tumor interactions [120]. Tumor-associated macrophages (TAMs) and stromal factors (VEGF, TGF-β) further enhance inflammation and cytokine release [121, 122]. Monitoring biomarkers such as serum IL-6, CRP, ferritin, and IFN-γ can help identify high-risk patients [123]. First-line management includes tocilizumab and corticosteroids, though these may reduce CAR-T persistence [124]. Emerging approaches involve CAR-T engineering with suicide switches or cytokine-neutralizing constructs (e.g., IL-6 “trap”) to mitigate CRS while preserving antitumor efficacy [117, 125]. Integrating such controls is essential as CAR-Ts evolve toward multi-antigen targeting and armored designs capable of modulating the suppressive TME without systemic hyperinflammation [126].

Hydrogel therapy (injectable hydrogel)

Hydrogels are three-dimensional, hydrophilic polymeric networks capable of absorbing substantial amounts of water while maintaining structural integrity [127]. Their physicochemical versatility, biocompatibility, and tunable mechanical properties have positioned them as attractive candidates for biomedical applications, particularly in tissue engineering, drug delivery, and immunotherapeutic platforms. Hydrogels can be classified based on their origin (natural vs. synthetic), polymeric composition, crosslinking mechanism (physical or chemical), and responsiveness to stimuli such as pH, temperature, or enzymatic activity. Natural hydrogels, derived from materials like alginate, collagen, hyaluronic acid, or chitosan, provide intrinsic bioactivity and degradability, while synthetic hydrogels such as polyethylene glycol (PEG), polyvinyl alcohol (PVA), and poloxamers afford more precise control over mechanical properties and structural customization [15].

Hydrogels are broadly categorized into conventional (non-responsive) and smart (stimuli-responsive) systems [19]. Conventional hydrogels function as passive scaffolds, support the encapsulation and slow release of immune cells or modulatory agents. In contrast, smart hydrogels are engineered to respond dynamically to environmental cues such as pH, temperature, enzymatic activity, or oxidative stress [15]. Furthermore, hydrogels can be functionalized with integrin-binding peptides (e.g., RGD), chemokines (e.g., CXCR4 inhibitor), or checkpoint inhibitors (e.g., anti-PD-1) to modulate immune cell trafficking, survival, and effector function at the implantation site [14, 16, 128–130].

Biophysical regulation of T-cell fate by hydrogel microenvironments

Accumulating evidence from immune mechanobiology demonstrates that T cells are not purely biochemically regulated immune cells, but rather function as mechanosensitive systems whose activation, differentiation, migration, and exhaustion states are tightly controlled by mechanotransductive signaling pathways. Mechanical forces, cytoskeletal dynamics, and physical constraints act in concert with canonical TCR signaling to determine T-cell fate [131].

Hydrogels, therefore, should be conceptualized not as passive scaffolds but as active immunoregulatory niches, whose biophysical properties directly shape T-cell phenotype and function.

Stiffness (mechanical cues) and T-cell phenotype regulation

T-cell activation is fundamentally a force-sensitive process. Productive signaling depends on mechanical tension generated at the TCR–pMHC interface, where applied forces promote conformational rearrangements that potentiate downstream cascades. This response is coordinated by cytoskeletal remodeling—including actin polymerization, myosin II–driven contractility, and integrin-mediated adhesion—which together constitute a mechanotransductive network coupling extracellular physical inputs to antigen recognition. Within this context, hydrogels serve as programmable biomaterial systems capable of modulating force transmission, cytoskeletal prestress, and activation thresholds. By shaping these biophysical parameters, they function not merely as structural supports but as active determinants of T-cell transcriptional and metabolic programs [132].

Hydrogels engineered to emulate the compliant mechanical properties of lymphoid tissues—defined by a low elastic modulus and physiologic viscoelastic behavior—establish microenvironments that are conducive to optimal T-cell function. In such mechanically permissive settings, actomyosin contractility remains appropriately regulated, TCR signaling is integrated with greater stability, and mechanotransductive activation occurs in a controlled manner. These conditions are associated with the preservation of naïve and central memory subsets, maintenance of stem-like memory characteristics, and enhanced metabolic adaptability. Importantly, precise modulation of crosslinking density, polymer composition, and stress-relaxation dynamics allows fine control over intracellular force transmission and downstream mechanosensitive signaling. Through this mechanical tunability, hydrogels can serve as instructive platforms that influence T-cell fate decisions independently of exogenous biochemical cues [133].

Conversely, hydrogels designed to replicate the rigidity of tumor or fibrotic tissues expose T cells to increased mechanical resistance, heightened integrin engagement, and prolonged cytoskeletal tension. These conditions drive sustained force-dependent TCR signaling, induce metabolic stress, disrupt calcium homeostasis, and trigger transcriptional programs associated with exhaustion. Such mechanopathological responses highlight the dual utility of hydrogels, serving both as experimental platforms and as immunoengineering tools capable of modeling physiological and pathological mechanical environments. By integrating spatially patterned stiffness gradients and tuning viscoelastic properties, these materials provide programmable control over mechanotransductive pathways, offering a strategy to limit T-cell exhaustion, maintain functional persistence, and enhance the long-term efficacy of immunotherapies.

Porosity, microarchitecture, and migration-phenotype coupling

The porosity and structural organization of three-dimensional microenvironments serve as key regulators of T-cell migration, mechanosensing, and phenotypic programming. Amoeboid T-cell motility is particularly sensitive to variations in pore size, spatial confinement, and matrix density, which influence cytoskeletal architecture, intracellular tension, and nuclear deformation, ultimately impacting transcriptional and metabolic states. Networks that are highly porous and well-interconnected alleviate mechanical constraints, facilitating rapid migration, effective immune surveillance, and reduced cytoskeletal stress. In contrast, dense and restrictive matrices impair actomyosin coordination, disrupt calcium signaling, and hinder cellular infiltration, fostering sustained mechanotransductive stress and the emergence of dysfunctional or exhaustion-prone phenotypes.

Hydrogels and other engineered biomaterials offer highly tunable platforms to investigate how physical and mechanical cues regulate immune cell behavior, providing precise control over porosity, pore connectivity, mechanical stiffness, and biochemical composition. In organ-on-a-chip and immune-on-a-chip systems, hydrogel architectures are designed to replicate physiologically relevant extracellular matrices, incorporating perfusable and porous networks that allow immune cells to navigate three-dimensional spaces while maintaining physiologic shear forces, confinement, and biochemical gradients. These engineered systems have shown that larger, interconnected pores enhance cell infiltration, spatial distribution, and network formation, whereas restrictive pore geometries can limit mechanosensitive migration and downstream signaling, thereby directly influencing immune cell phenotype and function in vitro. Increasingly, microfluidic immune models incorporate hydrogel ECMs to mimic interstitial pathways, guiding immune cell transmigration in response to chemotactic cues and highlighting porosity as a critical regulator of immune mobilization and phenotypic plasticity [134].

Microarchitectural properties of three-dimensional matrices play a central role in shaping mechanotransduction pathways within immune cells, linking physical confinement and matrix stiffness to intracellular signaling networks that govern differentiation, metabolic activity, and cellular persistence. Environments with high porosity and minimal confinement help reduce aberrant mechanosignaling, relieve prolonged actomyosin tension, and maintain balanced metabolic flux, supporting phenotypes associated with effective immune surveillance and memory formation. In contrast, dense or restrictive architectures increase mechanical load, perturb mechanosensitive transcriptional programs, and can accelerate exhaustion, ultimately compromising long-term functional resilience. By employing hydrogel systems with tunable porosity and viscoelasticity, researchers can precisely manipulate these mechanobiological cues independently of biochemical signals, enabling the rational design of immune-instructive niches that enhance migration, preserve effector function, and stabilize phenotypes for translational immunoengineering and therapeutic applications [135].

Degradation kinetics and dynamic mechanical remodeling

Dynamic degradation and mechanically adaptive remodeling represent key biophysical factors that regulate immune cell behavior within three-dimensional matrices. In contrast, static, non-degradable scaffolds impose persistent mechanical confinement, maintaining elevated cytoskeletal tension and aberrant mechanotransductive signaling—conditions closely linked to T-cell dysfunction and the development of exhaustion commonly observed in solid tumor microenvironments [136].

By contrast, dynamically degradable hydrogels—incorporating stimuli-responsive bonds, enzyme-cleavable linkages, or reversible crosslinks—enable adaptive remodeling of the matrix, progressive stress relaxation, and controlled modulation of mechanical resistance, closely recapitulating physiological extracellular matrix turnover and the plasticity of immune niches. These dynamic features facilitate mechanical release, allowing T cells and CAR-T cells to migrate gradually, alleviate prolonged intracellular tension, and avoid sustained activation signals that promote exhaustion. When combined with tunable viscoelastic properties, such hydrogels support adaptive locomotion, preserve functional phenotypes, and maintain immune competence in ways that static scaffolds cannot. Notably, dynamic hydrogels have emerged as versatile biomaterials capable of mimicking the evolving tumor microenvironment, enhancing immune cell infiltration, modulating local mechanical and biochemical cues, and improving the efficacy of immunotherapies by mitigating chronic mechanical stress that would otherwise impair effector function [137, 138].

Lymphoid tissue mimicry vs tumor biophysical mimicry

Lymphoid tissues are characterized by soft, porous, and dynamically remodeled extracellular matrices that naturally support T-cell trafficking, clonal expansion, memory differentiation, and long-term persistence. These microenvironments provide low mechanical resistance, physiologic stress relaxation, and permissive mechanotransductive cues, which together promote balanced cortical contractility and efficient interstitial migration while limiting chronic mechanical stress that can drive cellular dysfunction. Biomimetic platforms designed to replicate these biomechanical features—including hydrogels with tunable stiffness, porosity, and viscoelastic stress-relaxation properties—effectively preserve memory-associated phenotypes and reduce markers of exhaustion by recapitulating the essential biophysical characteristics of lymphoid niches. As a result, such engineered systems enhance T-cell functional resilience and migratory competence both in vitro and in advanced culture models [139].

Conversely, solid tumor microenvironments are characterized by increased stiffness, dense fibrotic architectures with enhanced collagen crosslinking, and limited dynamic remodeling. These features impose sustained physical confinement, disrupt cytoskeletal dynamics, and hinder CAR-T and T-cell infiltration, resulting in mechanosensitive suppression of migration and reduced effector function within three-dimensional matrices. Within such stiff, tumor-mimetic contexts, CAR-T cells display impaired infiltration and motility, reflecting the mechanotransductive barriers imposed by the matrix. These observations underscore the critical role of soft, lymphoid-mimicking hydrogel ECMs in maintaining immune competence and overcoming the physical constraints that limit T-cell function in solid tumors [136].

Combining hydrogel engineering and immunotherapy for advanced breast cancer treatment

Several preclinical studies have highlighted the advantages of hydrogel-encapsulated therapies in breast cancer models which demonstrate superior antitumor efficacy compared to conventional drug delivery. By way of example, a sodium alginate-based hydrogel loaded with nanoparticle albumin-bound paclitaxel (Nab-PTX) and the Toll-like receptor 7 (TLR7) agonist R837 induced robust immunogenic cell death in murine breast tumors. This combinatorial approach enhanced dendritic cell activation and simultaneously promoted the expansion of effector memory T cells which lead to significant tumor regression and prolonged survival. Notably, animals treated with the hydrogel formulation exhibited durable immune memory upon tumor rechallenge, underscoring the potential of localized hydrogel delivery to augment systemic antitumor immunity [140].

A recent study by Mantooth et al. has introduced an innovative injectable hydrogel platform, XCSgel, to improve cytokine immunotherapy outcomes in murine models of triple-negative breast cancer (TNBC). This hydrogel, composed of a chitosan-based crosslinked matrix, was employed to locally deliver interleukin-12 (IL-12) directly into the tumor site. The hydrogel significantly prolonged the intratumoral retention of IL-12, enhanced its therapeutic window, and minimized systemic exposure. In vivo experiments have demonstrated robust antitumor effects, with a single injection leading to complete regression in the majority of E0771 tumors and a subset of mWnt tumors. Remarkably, mice whose tumors were completely eliminated were able to resist the growth of new tumors when exposed again due to a strong and lasting immune memory response. Immunophenotyping further revealed a substantial increase in proliferative, activated CD8⁺ T cells and a marked reduction in exhausted CD8⁺ populations within the tumor microenvironment. These findings underscore the potential of hydrogel-facilitated cytokine delivery to reshape local immunity and induce lasting systemic protection against TNBC [141].

In 2021, Kim et al. demonstrated that hydrogel-based platforms can be utilized to synergize photodynamic therapy (PDT) with immunomodulation in breast cancer models. An intratumorally administered hyaluronic acid hydrogel loaded with the photosensitizer chlorin e6 (Ce6) and TLR7 agonist R837 demonstrated enhanced induction of immunogenic cell death following light activation. This triggered dendritic cell maturation and cytotoxic T lymphocyte activation result in both local tumor suppression and systemic antitumor immunity. Importantly, inhibition of primary tumor growth was accompanied by protection against tumor rechallenge indicate the hydrogel’s role in facilitating sustained immune stimulation and durable antitumor memory [142].

Zhang et al., in 2021, revealed that PEGylated hydrogels loaded with anti-PD-L1 antibodies significantly enhanced CD8⁺ T cell infiltration into tumors [143]. Similarly, Xu et al. (in 2024) reported that this approach effectively reduced regulatory T cell (Treg)-mediated immunosuppression within the tumor microenvironment. Together, these studies have highlighted the potential of PEGylated hydrogel-based delivery systems to improve antitumor immune responses by both promoting effector T cell activity and alleviating immunosuppressive mechanisms [129].

In a study, Gong et al. investigated the efficacy of umbilical cord blood natural killer (UCB-NK) cell-based immunotherapy when combined with hydrogel therapy, as compared to UCB-NK cell monotherapy. UCB-NK cell monotherapy is limited by the immunosuppressive tumor microenvironment which diminishes NK cell cytotoxicity through reduced MHC class I-related chain A/B (MICA/B) expression and autophagy-induced degradation of granzyme B (GZMB). To address this limitation, UCB-NK cells were co-delivered to the tumor site via an injectable hydrogel formulation containing suberoylanilide hydroxamic acid (SAHA), let-7e-5p, miR-615-3p, and 3-methyladenine (3-MA). SAHA modulates gene expression by promoting histone acetylation and influences key cellular processes such as apoptosis, cell cycle progression, autophagy, and differentiation. The microRNAs let-7e-5p and miR-615-3p function as tumor suppressors, while 3-MA inhibits autophagosome formation by blocking PI3K activity. This combinatorial strategy not only enhanced the cytotoxic function and persistence of UCB-NK cells but also supported hemostasis and accelerated wound healing, owing to the bioactive properties of the hydrogel matrix [144].

Harnessing hydrogel matrices to optimize CAR-T cell delivery, activation, and antitumor response

Hydrogels have emerged as a versatile delivery platform for immunotherapies, particularly in solid tumors as triple-negative breast cancer, where conventional systemic delivery often shows limited efficacy or fails to achieve sustained therapeutic benefit [145–148]. Engineered injectable hydrogels function as immunostimulatory depots that improve CAR-T cell persistence and prolong their antitumor cytotoxicity at the tumor site. Shear-thinning hydrogels allow minimally invasive intratumoral administration and help retain therapeutic agents at the tumor sites, creating a localized immunostimulatory environment rather that allowing rapid systemic clearance [8, 147]. Integration of immunomodulatory cues such as IL-15, IL-21, and checkpoint inhibitors (e.g., anti-PD-1), alongside chemokines like CXCR4 inhibitor, facilitates enhanced T cell recruitment, expansion, and persistence [149–151]. Furthermore, incorporating stimulus-responsive features can significantly improve the performance of hydrogels and broaden their biomedical applications. Smart hydrogels, a rapidly advancing class of materials, respond to external cues such as pH, temperature, light, electrical or magnetic fields, and biomolecule concentrations, enabling precise and controlled release of therapeutic cargo at targeted sites (Fig. 3) [148, 152]. This versatility allows hydrogels to be further customized using specific stimuli—including enzymatic activity, pH changes, or other tumor-associated signals—to achieve precise, localized release of therapeutic payloads, as exemplified by enzyme-responsive systems that selectively degrade within the tumor microenvironment. Mechanistically, these hydrogels are constructed using tumor-selective protease-cleavable peptide crosslinkers, such as MMP-2/9-sensitive motifs like the GPQG↓IAGQ sequence, which form integral components of the polymeric network. The selectivity arises from the careful design of peptide sequences that are specifically recognized and cleaved by enzymes active in the tumor microenvironment, including MMP-2 and MMP-9. Upon exposure to matrix metalloproteinases secreted by tumor and stromal cells, these peptide linkers undergo site-specific cleavage, resulting in localized network destabilization, increased mesh porosity, and gradual loss of mechanical integrity. This controlled remodeling of the hydrogel matrix enables spatially confined release of embedded CAR-T cells and guides their migration along chemokine gradients toward tumor sites. Importantly, the use of highly specific protease-responsive sequences and precise network-threshold degradation kinetics minimizes nonspecific hydrolysis and premature release, ensuring tumor-targeted activation and delivery of CAR-T cell therapeutics [153, 154].

Fig. 3.

Beyond serving as delivery platforms, hydrogels function as localized depots for cytokines and bioactive cues that enhance CAR-T cell recruitment, expansion, and persistence at the tumor site. Incorporation of immunostimulatory factors (e.g., IL-15, IL-21), checkpoint blockade (anti-PD-1), and chemokine modulation (e.g., CXCR4 inhibition) helps counteract T cell exhaustion and immunosuppressive signaling within the tumor microenvironment. Created in BioRender. shojaei, S. (2026) https://BioRender.com/bqsv1a8

These next-generation represent an optimal integration of materials science, synthetic immunology, and tumor biology, allowing precise control over immune cell programming and microenvironmental modulation. Beyond simply delivering cells, hydrogels are now being functioned as depots for cytokines and other bioactive factors, supporting and amplifying CAR-T cell activity at the tumor site [155]. These capabilities are crucial for overcoming T cell exhaustion and immunosuppressive signaling within the TME. Preclinical studies in murine breast cancer models have demonstrated that hydrogel-based CAR-T cell delivery significantly improves persistence, tumor infiltration, and on-target cytotoxicity compared to traditional systemic administration routes [156, 157]. Collectively, these innovations underscore the transformative potential of hydrogel-enabled CAR-T cell platforms in advancing solid tumor immunotherapy and set the stage for translational strategies in clinical oncology [158]. In the following sections, we explore how hydrogel-based strategies can be rationally engineered to overcome six critical barriers; namely, tumor microenvironment immunosuppression, poor infiltration and persistence, on-target/off-tumor toxicity and cytokine release syndrome (CRS). For each challenge, we examine the underlying immunopathology and present hydrogel-enabled innovations that reshape the tumor-immune interface, enhance CAR-T cell function, and improve clinical translatability (Table 2).

Table 2.

Material-specific hydrogel engineering strategies to overcome key CAR-T therapy challenges in solid tumors

CAR-T therapy challenge	Hydrogel material platform	Specific functional modification/design feature	Mechanistic rationale	Representative outcome	Representative reference
Immunosuppressive TME	Hyaluronic Acid (HA) hydrogel	Encapsulated anti-PD-L1 nanoparticles + IL-15 co-delivery	Local checkpoint blockade + sustained cytokine signaling enhances CAR-T persistence	↑ intratumoral CAR-T density, ↓ recurrence	Hu et al. [130]
	PEG-based hydrogel	TGF-β inhibitor–loaded microspheres	Neutralizes local immunosuppressive cytokines	↓ exhaustion markers (PD-1, TIM-3)	Grosskopf et al. [159]
	Alginate hydrogel	RGD-functionalization	Enhances integrin-mediated adhesion & survival	↑ IFN-γ secretion	Zhang et al. [160, 182]
	Self-assembling peptide hydrogel	Tunable stiffness (LN-mimetic niche)	Mechanotransduction-driven activation & expansion	↑ CAR expression, ↑ proliferation	Jie et al. [145]
Poor Tumor Infiltration	MMP-cleavable HA hydrogel	Protease-sensitive linker	Degrades in ECM-remodeling environment → controlled T-cell release	↑ tumor penetration depth	Li et al. [18]
	Chitosan-PEG injectable hydrogel	Tuned pore size & porosity	Supports migration & sustained local retention	↑ intratumoral cell persistence	Suraiya et al. [161]
	Macroporous cryogel scaffold	Interconnected large pores	Facilitates rapid trafficking & oxygen diffusion	↑ migration kinetics	Xu et al. [129]
	Collagen-mimetic hydrogel	ECM-like fiber architecture	Provides structural guidance cues	Improved infiltration distribution	Kim et al. [162]
Cytokine Release Syndrome (CRS)	In situ-forming PEG hydrogel	pH-responsive retention system	Spatial confinement in acidic TME	↓ systemic IL-6 peak	Cheng et al. [163]
	HA hydrogel	Sustained IL-15 release profile	Avoids cytokine bolus effect	↓ cytokine burst magnitude	Cheng et al. [163]
	HPMC-C12 hydrogel	Tunable cytokine release kinetics	Promotes memory phenotype without hyperactivation	↓ peak inflammatory cytokines	Yu et al. [164]
	Shear-thinning injectable hydrogel	Local CAR-T depot formation	Limits systemic dissemination	↓ systemic toxicity	Wang et al. [111, 165]
Limited Persistence / Memory Formation	PEG–heparin hydrogel	Lymph-node-mimetic stiffness	Enhances viral transduction & expansion ex vivo	↑ %CAR⁺ cells (~ 2 ×)	Castellote-Borrell et al. [158]
	Self-assembling peptide hydrogel	Adhesive ligand density optimization	Supports TSCM and central memory subsets	↑ CD62L⁺ CCR7⁺ phenotype	Zhang et al. [160, 182]
	HA + IL-15 hydrogel	Sustained cytokine microenvironment	Maintains long-term effector memory	↑ persistence duration	Zeng et al. [166]
	Alginate cryogel	3D macroporous expansion scaffold	Provides ex vivo expansion niche	↑ fold-expansion rate	Fatoni et al. [167]
Off-Tumor Toxicity / Spatial Control	PEG hydrogel	Acid-sensitive linker	Tumor-specific degradation	↓ healthy tissue damage	Cheng et al. [163]
	ROS-responsive hydrogel	Oxidative cleavage bonds	Releases cells preferentially in tumor oxidative stress	↑ safety index	Huang et al. [168]
	Thermosensitive PLGA-PEG hydrogel	Temperature-triggered gelation	Forms depot post-injection	↓ systemic exposure	Elhabal et al. [169]
	Photo-crosslinkable hydrogel	Spatially controlled polymerization post-resection	Precise localization in surgical cavity	↓ local recurrence	Xue et al. [170]

Reshaping the tumor microenvironment with hydrogel-enhanced CAR-T cell therapy

The immunosuppressive tumor microenvironment (TME) significantly restricts CAR-T cell efficacy in breast cancer, characterized by dense stroma, immunoregulatory cytokines (e.g., TGF-β, IL-10), myeloid-derived suppressor cells (MDSCs), and tumor-associated macrophages (TAMs) that collectively dampen effector T cell responses [171, 172]. One of the most significant effects of this suppressive environment is the development of T-cell exhaustion. In this state, T cells gradually lose their killing capacity, produce fewer cytokines, undergo metabolic disruption, and maintain high expression of inhibitory receptors such as PD-1, LAG-3, and TIM-3 [173, 174].

Hydrogels offer a highly adaptable platform to modulate the TME locally and prevent exhaustion through multiple synergistic mechanisms. First, the encapsulation of CAR-T cells within hydrogels co-loaded with TME-neutralizing agents, such as IDO inhibitors, CSF-1R antagonists, or TGF-β blockers, effectively silences local immunosuppressive signals that otherwise drive the upregulation of exhaustion markers and impair metabolic fitness [18, 175]. By normalizing the cytokine milieu, these hydrogels preserve CAR-T cell effector functions and promote sustained IL-2 and IFN-γ production. Second, the hydrogel matrix itself can be engineered to deliver checkpoint blockade antibodies (e.g., anti-PD-1, anti-CTLA-4) in a controlled and spatially confined manner, directly antagonizing exhaustion pathways in situ without inducing systemic immune-related adverse events [127, 176].

Moreover, many hydrogels are engineered to provide immunometabolic support, such as supplying interleukin-15 or mitochondrial protective agents, which help maintain oxidative phosphorylation and glycolytic capacity thereby promotes T-cell persistence; both of which are essential for effector T cell persistence under hypoxic, nutrient-deprived conditions [177, 178]. Some hydrogel designs also enable co-delivery of stimulatory cytokines (e.g., IL-21) or costimulatory ligands (e.g., 4-1BBL) that reinforce T cell activation thresholds and prevent the progression from transient dysfunction to irreversible exhaustion [179]. In addition, by remodeling the ECM using embedded enzymes like hyaluronidase or collagenase, hydrogels reduce physical barriers to T cell movement and restore immune surveillance [180, 181]. Taken together, these immunoengineered hydrogel systems create a spatially defined, proinflammatory niche that rescues CAR-T cells from exhaustion, supports their long-term viability, and transforms the TME from an immunosuppressive sink into an immunotherapeutic amplifier.

Building on this concept, Hu et al. [130] developed an injectable hyaluronic acid hydrogel system co-delivering CAR-T cells and anti-PD-L1-conjugated platelets to the tumor resection site. This platform enhanced local immune activation by improving CAR-T cell retention and reversing T cell exhaustion via PD-L1 blockade. In murine models, the strategy significantly suppressed post-surgical tumor recurrence and improved overall survival [130].

Improving CAR-T cell trafficking and cytotoxicity in breast cancer via hydrogel scaffolds

Effective trafficking and infiltration of CAR-T cells into solid tumors such as breast cancer remain a critical challenge, hindered by the tumor’s dense extracellular matrix (ECM), abnormal vasculature, and chemokine-receptor mismatches. Hydrogel scaffolds offer a rational design strategy to overcome these barriers by enhancing local delivery and sustained presence of CAR-T cells at tumor sites. Biocompatible hydrogels can be engineered with defined pore sizes and MMP-sensitive crosslinkers to allow CAR-T cell retention and tumor-responsive release [8, 157]. Incorporation of chemokines like CCL21 or CXCL9 within the hydrogel matrix creates chemotactic gradients that improve both CAR-T and host T cell homing toward tumor tissues [182]. Moreover, environment-responsive degradation tailored to the tumor’s proteolytic activity ensures that CAR-T cells are released at the optimal location and time, enabling enhanced infiltration into tumor masses and more effective engagement with malignant cells [183].

Beyond trafficking, hydrogel scaffolds are being explored as local immunomodulatory depots that enhance CAR-T cytotoxic function in situ. By embedding cytokines such as IL-15 or IL-21 into the matrix, these platforms promote CAR-T cell expansion, enhance expression of effector molecules like granzyme B and perforin, and delay T cell exhaustion [145, 184]. In parallel, local delivery of immune checkpoint inhibitors (e.g., anti-PD-1) or metabolic modulators (e.g., IDO or arginase inhibitors) within the hydrogel enhances CAR-T activity by counteracting the suppressive breast cancer TME [185]. Such localized immune engineering not only reduces systemic side effects, including cytokine release syndrome, but also increases tumor-specific cytotoxicity. In murine models of triple-negative breast cancer, hydrogel-assisted CAR-T therapies have demonstrated improved intratumoral retention, enhanced tumor lysis, and prolonged survival compared to systemic administration routes [186, 187]. Collectively, hydrogel scaffolds represent a transformative approach to co-optimize CAR-T cell trafficking and function for more durable and localized antitumor immunity in breast cancer.

Extending this line of research, in a recent study, Suraiya et al. developed micro-scale injectable hydrogels capable of locally delivering CAR-T cells to solid tumor spheroids and effectively enhancing T cell–tumor contact and infiltration. This platform provided a 3D matrix that mimicked the tumor microenvironment which support CAR-T cell viability, expansion, and cytotoxicity. Compared to free CAR-T cell administration, the hydrogel formulation significantly enhanced cytotoxicity and reduced immune escape, while maintain CAR-T cell functionality within the immunosuppressive tumor core [161].

Similarly, Li et al. [18] engineered a photocurable hydrogel-based delivery system for the co-administration of CAR-T cells and the mitophagy agonist BC1618 to enhance antitumor responses in triple-negative breast cancer. The hydrogel, composed of a light-activated polymer matrix and photoinitiator, was injected into the tumor site and solidified in situ under 405 nm violet light, forming a three-dimensional depot that retained both CAR-T cells and BC1618 within the tumor microenvironment. This system enabled localized, sustained release, ensuring prolonged exposure of CAR-T cells to the mitophagy agonist and preventing their systemic dilution or rapid clearance. Functionally, the hydrogel provided a structural niche that supported CAR-T cell viability, infiltration, and expansion, while the mitophagy agonist enhanced mitochondrial fitness and reduced exhaustion by promoting immunogenic cell death. Consequently, this combinatorial approach led to significant tumor regression, reduced T cell exhaustion, and increased cytokine secretion [18].

In a related approach, Chao et al., in a separate study, achieved comparable outcomes by incorporating metformin into the hydrogel, which reduced oxidase activity of cancer cell in TME [155].

Smart hydrogel systems for precision-controlled CAR-T cell expansion and targeted breast cancer therapy

Effective clinical translation of CAR-T cell therapies for solid tumors like breast cancer requires modular platforms that can meticulously regulate CAR-T cell localization, activation, and expansion. Injectable, human-compatible hydrogels enable direct intratumoral or perioperative deposition of CAR-T cells, enhancing retention and minimizing systemic cytokine toxicity compared to intravenous delivery routes. These platforms can be tailored with controlled release kinetics to deliver cells over days to weeks, ensuring sustained exposure aligned with tumor clearance needs. Moreover, their injection-friendly formats are easily matched to existing clinical workflows, including minimally invasive tumor access and post-resection cavity filling, making them well-suited for real-world oncology settings [159].

Recent hydrogel systems extend this approach by enabling precise local release of CAR-T cells and real-time monitoring of their activity. These systems support pulsatile or sustained cell release, maintaining effective CAR-T cell concentrations within tumors while reducing off-target exposure and clearance [82, 188]. Integrating biosensors, imaging contrast agents, or molecular diagnostics within the hydrogel matrix enables noninvasive tracking of CAR-T cell proliferation, activation, and localization—facilitating adaptive treatments based on direct feedback [189]. Additionally, these reservoirs can co-deliver cytokines or chemokines (such as IL-15), creating pro-inflammatory niches that amplify CAR-T expansion and engage local immunity [179, 190]. By merging controlled delivery, immune augmentation, and smart sensing into a single construct, these next-generation hydrogels provide a multidimensional solution to the challenges of solid tumor CAR-T therapy, charting a path toward personalized therapeutic strategies in metastatic breast cancer.

In support of this strategy, Stephan et al. [191] demonstrated that synthetic stimulatory molecules can be bound to T cells or embedded within hydrogels to mimic key features of the immunological synapse, thereby promoting more potent and sustained T cell activation [191].

Enhancing tumor specificity to prevent off-tumor toxicity

By integrating site-specific delivery, immune potentiation, and environmental responsiveness, hydrogel constructs offer a precision-targeted strategy for CAR-T cell therapy that limits systemic trafficking and markedly reduces on-target/off-tumor toxicity in solid tumors [192, 193].

For instance, injectable polymer–nanoparticle (PNP) hydrogels, composed of dodecyl-modified hydroxypropyl methylcellulose (HPMC-C₁₂) and PEG–PLA nanoparticles, establish a transient inflammatory niche that retains CAR-T cells at the tumor site. These hydrogels enable sustained T cell activation through co-delivered cytokines and restrict systemic dissemination, thereby minimizing exposure to antigen-positive healthy tissues while enhancing tumor infiltration and therapeutic efficacy in murine solid tumor models [159].

Similarly, hyaluronic acid-based hydrogels co-loaded with CAR-T cells, IL-15 nanoparticles, and PD-L1–conjugated platelets have demonstrated the ability to form localized immune reservoirs within surgical resection cavities. These systems support regional immune activation, prevent tumor recurrence, and limit systemic CAR-T cell dispersion, offering a clinically promising approach to mitigate off-target cytotoxicity [130].

As a further example, Song et al. [194] developed a sprayable hydrogel composed of sulfated hyaluronic acid, chitosan, and F127 for post-surgical adhesion prevention. Beyond its mechanical barrier function, this hydrogel exhibited robust immunomodulatory activity,in a rat peritoneal injury model, it reprogrammed pro-inflammatory M1 macrophages toward an anti-inflammatory M2 phenotype via CD44 receptor engagement and downstream STAT6 activation. This phenotype shift disrupted macrophage clustering and limited local fibrosis by attenuating NF-κB-driven cytokine release. Importantly, this study illustrates how engineered hydrogels can precisely modulate innate immune behavior at specific anatomical sites; offering a conceptual framework for mitigating on-target/off-tumor toxicity in CAR-T cell therapies. By confining immune activity through local hydrogel application and skewing inflammatory responses toward tissue-preserving phenotypes, similar platforms may reduce collateral damage in healthy tissues expressing low levels of tumor-associated antigens, thus enhancing the safety profile of CAR-T treatment in solid tumors [194].

Strategic modulation of CAR-T cell activation to mitigate cytokine release syndrome (CRS)

Cytokine Release Syndrome (CRS) represents a major dose-limiting and life-threatening toxicity associated with CAR-T cell therapies, particularly in hematologic and increasingly in solid tumors. CRS is initiated by supraphysiologic activation of infused CAR-T cells upon encountering tumor-associated antigens, leading to the release of a storm of proinflammatory cytokines, including IL-6, IFN-γ, TNF-α, and GM-CSF [100, 118, 120]. These cytokines activate bystander immune cells (monocytes, macrophages, endothelial cells) that further amplify systemic inflammation through a feed-forward loop. The resulting capillary leakage, hypotension, and multiorgan dysfunction can be fatal if not promptly mitigated. While IL-6 receptor blockade (e.g., tocilizumab) is the current mainstay for CRS management, it acts downstream of the initial immune cascade and does not address the source of cytokine hypersecretion; namely, the uncontrolled activation and proliferation of CAR-T cells [124].

Notably, the safety benefit of hydrogel-based CAR-T cell delivery arises primarily from the spatiotemporal regulation of T-cell activation and migration, rather than from passive cytokine sequestration. By locally confining CAR-T cells, these hydrogels limit early systemic dispersion, modulate the kinetics of antigen exposure, and restrict excessive clonal expansion, thereby mitigating the initial cytokine surge that drives CRS. Although some hydrogel matrices may additionally retain or adsorb cytokines through physicochemical interactions, this effect serves as a supplementary rather than primary mechanism for controlling CRS.

Emerging hydrogel-based delivery systems provide a versatile strategy to precisely regulate CAR-T cell activation and spatial distribution, reducing the risk of CRS while maintaining therapeutic efficacy [195]. By restricting CAR-T cells within matrices adjacent to tumors, hydrogels provide spatial regulation of antigen exposure and control over T-cell proliferation [196]. Moreover, bioresponsive hydrogels engineered with immunoregulatory or metabolic modulators—such as PD-L1 mimetics, IDO-inducing agents, or hypoxia-sensitive release mechanisms—can help restrain CAR-T cell overactivation during the early post-infusion period [117, 197]. These platforms can be engineered to trigger T-cell release in response to localized tumor microenvironment signals—such as acidic pH or matrix metalloproteinase activity—thereby limiting systemic exposure to the initial surge of inflammatory mediators [198]. Additionally, co-encapsulating anti-inflammatory factors—such as IL-10 or TGF-β modulators—or regulatory immune cells can further protect surrounding tissues from collateral cytokine-mediated damage while promoting tolerogenic reprogramming of the innate immune compartment [126, 127].

From bench to clinic: hydrogel platforms for clinical CAR-T delivery in breast cancer

Engineering scalable, injectable hydrogel systems

Transformational hydrogel systems aim to bridge lab innovation and clinical reality by emphasizing good manufacturing practice (GMP)-compatible materials, injection-friendly formats, and seamless integration with surgical oncology workflows [199]. Biocompatible, injectable hydrogels (e.g., fibrin, collagen, hyaluronic acid, alginate) are being adapted for intratumoral or post-operative tumor-cavity infusion. These materials offer stable, scalable manufacturing pathways and controlled release kinetics that limit systemic exposure and toxicity while preserving CAR-T viability and function in situ [161, 200]. For instance, a hyaluronic acid hydrogel releasing CAR-T cells, IL-15 nanoparticles, and anti-PD-L1–conjugated platelets prevented post-surgical recurrence and stimulated abscopal immunity in preclinical melanoma models, providing a strong translational precedent for breast cancer applications [130, 201].

Beyond functional delivery, advanced hydrogel designs embed real-time monitoring tools, such as imaging probes, biosensors, or release-reporting chemistries, enabling dynamic tracking of CAR-T cell localization, expansion, and activation [202]. These systems support adaptive therapeutic strategies and early detection of safety signals, complementing current efforts to mitigate CRS and off-target toxicity. Injectable micro-hydrogels harboring chemotactic molecules and oxygen-modulating drugs have enhanced CAR-T infiltration and cytotoxicity in solid tumors [203]. As regional therapies gain momentum in trials, these intelligent scaffolds could evolve into next-generation therapeutic reservoirs that allow clinicians to dynamically modulate cell dosing, monitor immune status, and optimize patient response; marking a significant step toward personalized CAR-T immunotherapy in breast oncology [52, 69].

Hydrogel-based CAR-T cell delivery for metastatic breast cancer

Metastatic breast cancer presents unique therapeutic challenges due to the dissemination of tumor cells across multiple organs and the heterogeneity of tumor microenvironments [25, 171]. Conventional systemic administration of CAR-T cells often results in poor homing efficiency, rapid clearance, and unintended off-target effects, limiting their clinical efficacy [148, 203]. Injectable hydrogel delivery platforms provide a promising alternative by enabling localized, site-specific deposition of CAR-T cells within metastatic lesions or their microenvironment. These biocompatible and tunable hydrogels conform to irregular tissue architectures and sustain CAR-T cell retention over extended durations, enhancing local cell density and functional persistence at metastatic sites [19, 141]. Moreover, hydrogels engineered with stimuli-responsive features, such as enzymatic degradation or pH-triggered release, allow precise CAR-T cell liberation in response to metastatic niche-specific biochemical cues, maximizing antitumor activity while minimizing systemic toxicity [204, 205].

Beyond serving as cellular reservoirs, hydrogel scaffolds actively reshape the immunosuppressive milieu of metastatic breast cancer. By co-delivering immunostimulatory molecules, including cytokines like IL-15 or checkpoint blockade antibodies, these platforms potentiate CAR-T cell cytotoxicity and counteract stromal fibrosis, hypoxia, and immune escape mechanisms typical of metastatic sites [179, 206]. The porous architecture facilitates infiltration and retention of both transferred and endogenous immune cells, fostering synergistic antitumor immunity [18]. Integration of diagnostic modalities, such as embedded biosensors or imaging contrast agents, further enables real-time monitoring of CAR-T cell kinetics, viability, and therapeutic response [188, 189]. In aggregate, hydrogel-based delivery systems offer a multifaceted, adaptable strategy to overcome biological and clinical barriers in metastatic breast cancer, advancing the promise of personalized, effective CAR-T cell immunotherapy (Table 2).

Manufacturing and regulatory considerations for injectable cell-laden hydrogels

Although injectable biomaterial systems provide significant advantages for localized delivery and immunomodulation, their translation into clinically viable products presents considerable manufacturing and regulatory challenges. In practice, hydrogels must either be sterilized before cell incorporation or produced entirely under aseptic conditions, since post-encapsulation sterilization is incompatible with living cells and bioactive molecules. Consequently, the clinical development of cell-laden injectable systems requires careful consideration of three critical domains to ensure safety, efficacy, and regulatory compliance.

Sterilization and aseptic manufacturing

Conventional terminal sterilization methods, including autoclaving, gamma irradiation, and ethylene oxide treatment, although effective for acellular biomaterials, can compromise cellular membranes and protein integrity. As a result, clinically relevant approaches rely on pre-sterilizing hydrogel precursors—such as through sterile filtration or irradiation of acellular components—followed by aseptic formulation and cell encapsulation under GMP-compliant cleanroom conditions. This strategy shifts the focus of sterility assurance from terminal sterilization to rigorously validated sterile processing workflows, ensuring microbial safety while maintaining CAR-T cell viability and preserving the functionality of bioactive proteins [207].

Rheological protection during injection

Injectable hydrogels must be carefully designed to minimize mechanical stress on CAR-T cells during delivery through narrow-gauge needles. Excessive shear forces can compromise cell membranes, disrupt cytoskeletal organization, and impair cellular function. To address this, hydrogels should exhibit shear-thinning properties, allowing viscosity to decrease under applied stress and rapidly recover after injection. Materials built on dynamic, reversible crosslinking networks effectively dissipate mechanical forces, providing a protective microenvironment that maintains cellular integrity, viability, and therapeutic functionality throughout the delivery process [208]. Optimizing these hydrogels requires a careful balance between injectability and mechanical stability after implantation, typically evaluated through rheological analyses at shear rates relevant to clinical delivery via narrow-gauge needles (e.g., 25–30 G). Demonstrating that cell viability and functional activity are preserved under clinically relevant injection conditions is critical to support translational potential and satisfy regulatory standards.

Clinical-grade production and regulatory complexity

Cell-laden hydrogels are classified as cell–material combination products and therefore demand integrated GMP-compliant manufacturing, multi-tiered quality control, and coordinated regulatory oversight. Key challenges include scalable production, ensuring batch-to-batch consistency, maintaining sterility, performing lot release testing for both cellular and material components, and addressing limited shelf life. These considerations underscore that successful clinical translation relies not only on biomaterial innovation but also on robust regulatory strategies and well-established manufacturing infrastructure [209].

Conclusion

In conclusion, these findings illustrate that hydrogel-T cell combination strategies represent a rational and powerful approach for treating breast cancer. By locally modulating the immune microenvironment and synchronizing the delivery of both cells and adjuvants, hydrogel systems can enhance T cell infiltration, activation, and persistence within the tumor bed [210]. As these technologies evolve, their tunability and biocompatibility make them ideal candidates for clinical translation in combination immunotherapies for breast cancer and beyond. Despite promising results, several hurdles remain in translating hydrogel-based immunotherapies into routine clinical practice. These include scalable manufacturing, batch consistency, and ensuring hydrogel degradation aligns with therapeutic timelines. Moreover, the complexity of breast tumor immunophenotypes demands personalized hydrogel formulations, potentially involving patient-derived biomarkers [211]. Future innovations may integrate bioresponsive hydrogels with real-time imaging, CRISPR-mediated immune editing, or synergistic modalities like radiotherapy. Taken together, these findings establish hydrogel therapy as a transformative frontier in breast cancer immunotherapy, with the potential to enhance therapeutic specificity, durability, and patient outcomes [212].

Abbreviations

BC

Breast cancer

TNBC

Triple-negative breast cancer

CAR-T cell

Chimeric antigen receptor T cell

DC

Dendritic cell

IL

Interleukin

PD-1

Programmed cell death protein 1

PD-L1

Programmed death-ligand 1

ECM

Extracellular matrix

TME

Tumor microenvironment

TNF-α

Tumor necrosis factor alpha

UCB-NK

Umbilical cord blood–derived natural killer (cell)

VEGF

Vascular endothelial growth factor

CAF

Cancer-associated fibroblast

3-MA

3-Methyladenine

4-1BB

CD137 (Tumor necrosis factor receptor superfamily member 9)

CCR2b

C–C chemokine receptor type 2b

Ce6

Chlorin e6

CRP

C-reactive protein

HER2

Human epidermal growth factor receptor 2

CRS

Cytokine release syndrome

CTL

Cytotoxic T lymphocyte

CXCL

Chemokine (C–X–C motif) ligand

CXCR

Chemokine (C–X–C motif) receptor

EGFR

Epidermal growth factor receptor

GM-CSF

Granulocyte–macrophage colony-stimulating factor

GMP

Good Manufacturing Practice

GZMB

Granzyme B

HIF

Hypoxia-inducible factor

HPMC-C₁₂

Dodecyl-modified hydroxypropyl methylcellulose

IDO

Indoleamine 2,3-dioxygenase

IFN-γ

Interferon gamma

LAG-3

Lymphocyte activation gene 3

MDSC

Myeloid-derived suppressor cell

MHC

Major histocompatibility complex

MICA/B

MHC class I–related chain A/B

MMP

Matrix metalloproteinase

MPC

Mitochondrial pyruvate carrier

MUC1

Mucin 1

NK

Natural killer (cell)

NKT

Natural killer T (cell)

OXPHOS

Oxidative phosphorylation

PEG

Polyethylene glycol

PEG–PLA

Polyethylene glycol–polylactic acid

PNP

Polymer–nanoparticle

PVA

Polyvinyl alcohol

RGD

Arginine–glycine–aspartic acid (integrin-binding motif)

SAHA

Suberoylanilide hydroxamic acid

SDF-1α

Stromal cell–derived factor 1 alpha

STAT6

Signal transducer and activator of transcription 6

TAAs

Tumor-associated antigens

TAM

Tumor-associated macrophage

TGF-β

Transforming growth factor beta

TIM-3

 T-cell immunoglobulin and mucin-domain containing protein 3

TLR

Toll-like receptor

Author contributions

F.S. designed the review study and S. S., M. S., and M. L. contributed to collecting the data and writing the manuscript. N. A. contributed to editing the text. All authors have confirmed the final version of the manuscript and are accountable for the contents of all parts of the work. All the authors have read and approved the final manuscript.

Funding

This study was not financially supported by any institution or funding agency.

Data availability

All data are available in this article.

Declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.